Structural Basis for Nucleotide Binding and Reaction Catalysis in Mevalonate Diphosphate Decarboxylase

Energy Technology Data Exchange (ETDEWEB)

Barta, Michael L.; McWhorter, William J.; Miziorko, Henry M.; Geisbrecht, Brian V. (UMKC)

2012-09-17

Mevalonate diphosphate decarboxylase (MDD) catalyzes the final step of the mevalonate pathway, the Mg{sup 2+}-ATP dependent decarboxylation of mevalonate 5-diphosphate (MVAPP), producing isopentenyl diphosphate (IPP). Synthesis of IPP, an isoprenoid precursor molecule that is a critical intermediate in peptidoglycan and polyisoprenoid biosynthesis, is essential in Gram-positive bacteria (e.g., Staphylococcus, Streptococcus, and Enterococcus spp.), and thus the enzymes of the mevalonate pathway are ideal antimicrobial targets. MDD belongs to the GHMP superfamily of metabolite kinases that have been extensively studied for the past 50 years, yet the crystallization of GHMP kinase ternary complexes has proven to be difficult. To further our understanding of the catalytic mechanism of GHMP kinases with the purpose of developing broad spectrum antimicrobial agents that target the substrate and nucleotide binding sites, we report the crystal structures of wild-type and mutant (S192A and D283A) ternary complexes of Staphylococcus epidermidis MDD. Comparison of apo, MVAPP-bound, and ternary complex wild-type MDD provides structural information about the mode of substrate binding and the catalytic mechanism. Structural characterization of ternary complexes of catalytically deficient MDD S192A and D283A (k{sub cat} decreased 10{sup 3}- and 10{sup 5}-fold, respectively) provides insight into MDD function. The carboxylate side chain of invariant Asp{sup 283} functions as a catalytic base and is essential for the proper orientation of the MVAPP C3-hydroxyl group within the active site funnel. Several MDD amino acids within the conserved phosphate binding loop ('P-loop') provide key interactions, stabilizing the nucleotide triphosphoryl moiety. The crystal structures presented here provide a useful foundation for structure-based drug design.

Crystallization and preliminary X-ray diffraction analysis of mevalonate kinase from Methanosarcina mazei

International Nuclear Information System (INIS)

Zhuang, Ningning; Seo, Kyung Hye; Chen, Cong; Zhou, Jia; Kim, Seon Won; Lee, Kon Ho

2012-01-01

Recombinant mevalonate kinase from M. mazei has been crystallized. Diffraction data were collected to 2.08 Å resolution. Mevalonate kinase (MVK), which plays an important role in catalysing the biosynthesis of isoprenoid compounds derived from the mevalonate pathway, transforms mevalonate to 5-phosphomevalonate using ATP as a cofactor. Mevalonate kinase from Methanosarcina mazei (MmMVK) was expressed in Escherichia coli, purified and crystallized for structural analysis. Diffraction-quality crystals of MmMVK were obtained by the vapour-diffusion method using 0.32 M MgCl 2 , 0.08 M bis-tris pH 5.5, 16%(w/v) PEG 3350. The crystals belonged to space group P2 1 2 1 2, with unit-cell parameters a = 97.11, b = 135.92, c = 46.03 Å. Diffraction data were collected to 2.08 Å resolution

The mevalonate pathway in neurons: It's not just about cholesterol.

Science.gov (United States)

Moutinho, Miguel; Nunes, Maria João; Rodrigues, Elsa

2017-11-01

Cholesterol homeostasis greatly impacts neuronal function due to the essential role of this sterol in the brain. The mevalonate (MVA) pathway leads to the synthesis of cholesterol, but also supplies cells with many other intermediary molecules crucial for neuronal function. Compelling evidence point to a model in which neurons shutdown cholesterol synthesis, and rely on a shuttle derived from astrocytes to meet their cholesterol needs. Nevertheless, several reports suggest that neurons maintain the MVA pathway active, even with sustained cholesterol supply by astrocytes. Hence, in this review we focus not on cholesterol production, but rather on the role of the MVA pathway in the synthesis of particular intermediaries, namely isoprenoids, and on their role on neuronal function. Isoprenoids act as anchors for membrane association, after being covalently bound to proteins, such as most of the small guanosine triphosphate-binding proteins, which are critical to neuronal cell function. Based on literature, on our own results, and on the analysis of public transcriptomics databases, we raise the idea that in neurons there is a shift of the MVA pathway towards the non-sterol branch, responsible for isoprenoid synthesis, in detriment to post-squalene branch, and that this is ultimately essential for synaptic activity. Nevertheless new tools that facilitate imaging and the biochemical characterization and quantification of the prenylome in neurons and astrocytes are needed to understand the regulation of isoprenoid production and protein prenylation in the brain, and to analyze its differences on diverse physiological or pathological conditions, such as aging and neurodegenerative states. Copyright © 2017 Elsevier Inc. All rights reserved.

Analysis of acyl CoA ester intermediates of the mevalonate pathway in Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Seker, Tamay; Møller, Kasper; Nielsen, Jens

2005-01-01

The mevalonate pathway plays an important role in providing the cell with a number of essential precursors for the synthesis of biomass constituents. With respect to their chemical structure, the metabolites of this pathway can be divided into two groups: acyl esters [acetoacetyl CoA, acetyl Co......A, hydroxymethylglutaryl (HMG) CoA] and phosphorylated metabolites (isopentenyl pyrophosphate, dimethylallyl pyrophosphate, geranyl pyrophosphate, farnesyl pyrophosphate). In this study, we developed a method for the precise analysis of the intracellular concentration of acetoacetyl CoA, acetyl CoA and HMG CoA; and we...... used this method for quantification of these metabolites in Saccharomyces cerevisiae, both during batch growth on glucose and on galactose and in glucose-limited chemostat cultures operated at three different dilution rates. The level of the metabolites changed depending on the growth phase...

Overexpressing 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR in the lactococcal mevalonate pathway for heterologous plant sesquiterpene production.

Directory of Open Access Journals (Sweden)

Adelene Ai-Lian Song

Full Text Available Isoprenoids are a large and diverse group of metabolites with interesting properties such as flavour, fragrance and therapeutic properties. They are produced via two pathways, the mevalonate pathway or the 2-C-methyl-D-erythritol-4-phosphate (MEP pathway. While plants are the richest source of isoprenoids, they are not the most efficient producers. Escherichia coli and yeasts have been extensively studied as heterologous hosts for plant isoprenoids production. In the current study, we describe the usage of the food grade Lactococcus lactis as a potential heterologous host for the production of sesquiterpenes from a local herbaceous Malaysian plant, Persicaria minor (synonym Polygonum minus. A sesquiterpene synthase gene from P. minor was successfully cloned and expressed in L. lactis. The expressed protein was identified to be a β-sesquiphellandrene synthase as it was demonstrated to be functional in producing β-sesquiphellandrene at 85.4% of the total sesquiterpenes produced based on in vitro enzymatic assays. The recombinant L. lactis strain developed in this study was also capable of producing β-sesquiphellandrene in vivo without exogenous substrates supplementation. In addition, overexpression of the strain's endogenous 3-hydroxy-3-methylglutaryl coenzyme-A reductase (HMGR, an established rate-limiting enzyme in the eukaryotic mevalonate pathway, increased the production level of β-sesquiphellandrene by 1.25-1.60 fold. The highest amount achieved was 33 nM at 2 h post-induction.

Utilization of biodiesel by-product as substrate for high-production of β-farnesene via relatively balanced mevalonate pathway in Escherichia coli.

Science.gov (United States)

You, Shengping; Yin, Qingdian; Zhang, Jianye; Zhang, Chengyu; Qi, Wei; Gao, Lan; Tao, Zhiping; Su, Rongxin; He, Zhimin

2017-11-01

Farnesene has been identified as suitable jet fuel substitutes and metabolic engineering for microbial production of farnesene is an alternative and attractive route. In this study, due to accumulation of toxic intermediate isopentenyl pyrophosphate (IPP), an engineered Escherichia coli strain harboring heterologous mevalonate pathway produced only 4.11mg/L β-farnesene. Through higher-level expression of isopentenyl diphosphate isomerase and farnesyl diphosphate synthase to minimize the accumulated IPP, another engineered strain with relatively balanced mevalonate pathway was constructed and had the highest production of β-farnesene to date (8.74g/L) by Escherichia coli in a lab bioreactor. Furthermore, this is the first report on utilization of biodiesel by-product (simple purification) as substrate for high-production of β-farnesene by the engineered strain optimized and β-farnesene concentration reached 2.83g/L in a lab bioreactor. Therefore, the engineered strain optimized could be used as a platform host for high-production of other terpenoids using biodiesel by-product as substrate. Copyright © 2017 Elsevier Ltd. All rights reserved.

Manipulation of isoprenoid biosynthesis as a possible therapeutic option in mevalonate kinase deficiency

NARCIS (Netherlands)

Schneiders, Marit S.; Houten, Sander M.; Turkenburg, Marjolein; Wanders, Ronald J. A.; Waterham, Hans R.

2006-01-01

OBJECTIVE: In cells from patients with the autoinflammatory disorder mevalonate kinase (MK) deficiency, which includes the hyperimmunoglobulin D with periodic fever syndrome, MK becomes the rate-limiting enzyme in the isoprenoid biosynthesis pathway. This suggests that up-regulation of residual MK

Synthesis of the mevalonic acid labelled with "1"4C, "1"3C and "3H

International Nuclear Information System (INIS)

Rousseau, Bernard

1982-01-01

This thesis describes five new methods of synthesis of the (R,S) mevalonic acid adapted to the labelling with "1"4C and "1"3C in positions 4,5 or 5 or 3', or with tritium in position 3'. Three of them use the tri-oxa-2,4,10 adamantyl group as masked carboxyl function. The two others take benefit from the regioselectivity of the bis-hydro-boration of terminal acetylenics by the 9-borabicyclo [3-3-1]nonane. The acylation of the bis-trimethylsilyl lithiomalonate, and the chemistry of dithiannes are also involved. Acetylene and methyl iodide labelled with isotopes are used as cheap base products [fr

Mevalonate kinase deficiency: Evidence for a phenotypic continuum

NARCIS (Netherlands)

Simon, A; Kremer, H P H; Wevers, R A; Scheffer, H; de Jong, J G; van der Meer, J W M; Drenth, J. P.

2004-01-01

Both mevalonic aciduria, characterized by psychomotor retardation, cerebellar ataxia, recurrent fever attacks, and death in early childhood, and hyper-immunoglobulin D (hyper-IgD) syndrome, with recurrent fever attacks without neurologic symptoms, are caused by a functional deficiency of mevalonate

Hyper-IgD syndrome or mevalonate kinase deficiency.

NARCIS (Netherlands)

Stoffels, M.; Simon, A.

2011-01-01

PURPOSE OF REVIEW: The hyper-IgD and periodic fever syndrome (HIDS) is one of the classical monogenetic hereditary autoinflammatory disorders, and together with the more severe mevalonic aciduria it is also known as 'mevalonate kinase deficiency' (MKD). In this study, we will give an overview of the

Biosynthesis of sterols from mevalonate in a starfish, Coscinasterias acutispina

International Nuclear Information System (INIS)

Teshima, Shin-ichi; Kanazawa, Akio

1976-01-01

This study deals with the biosynthesis of sterols from mevalonate in a starfish, Coscinasterias acutispina. After injection of mevalonate-2- 14 C, the metabolites were investigated by using thin-layer, column, and gas-liquid chromatographic techniques. The detailed investigation of radioactive desmethylsterols showed that radioactivity was mainly associated with cholest-7-enol. However, there was no evidence for the incorporation of mevalonate-2- 14 C into C 26 -, C 28 -, and C 29 -sterols besides cholestanol and cholesterol. The results indicated that the starfish, C. acutispina, is capable of synthesizing at least cholest-7-enol from mevalonate via probably squalene and lanosterol etc. But not sterols other than C 27 -sterols. Also, it was suggested that the conversion of cholest-7-enol to cholesterol may not proceed in this starfish. (auth.)

Mevalonate Pathway Antagonist Suppresses Formation of Serous Tubal Intraepithelial Carcinoma and Ovarian Carcinoma in Mouse Models.

Science.gov (United States)

Kobayashi, Yusuke; Kashima, Hiroyasu; Wu, Ren-Chin; Jung, Jin-Gyoung; Kuan, Jen-Chun; Gu, Jinghua; Xuan, Jianhua; Sokoll, Lori; Visvanathan, Kala; Shih, Ie-Ming; Wang, Tian-Li

2015-10-15

Statins are among the most frequently prescribed drugs because of their efficacy and low toxicity in treating hypercholesterolemia. Recently, statins have been reported to inhibit the proliferative activity of cancer cells, especially those with TP53 mutations. Because TP53 mutations occur in almost all ovarian high-grade serous carcinoma (HGSC), we determined whether statins suppressed tumor growth in animal models of ovarian cancer. Two ovarian cancer mouse models were used. The first one was a genetically engineered model, mogp-TAg, in which the promoter of oviduct glycoprotein-1 was used to drive the expression of SV40 T-antigen in gynecologic tissues. These mice spontaneously developed serous tubal intraepithelial carcinomas (STICs), which are known as ovarian cancer precursor lesions. The second model was a xenograft tumor model in which human ovarian cancer cells were inoculated into immunocompromised mice. Mice in both models were treated with lovastatin, and effects on tumor growth were monitored. The molecular mechanisms underlying the antitumor effects of lovastatin were also investigated. Lovastatin significantly reduced the development of STICs in mogp-TAg mice and inhibited ovarian tumor growth in the mouse xenograft model. Knockdown of prenylation enzymes in the mevalonate pathway recapitulated the lovastatin-induced antiproliferative phenotype. Transcriptome analysis indicated that lovastatin affected the expression of genes associated with DNA replication, Rho/PLC signaling, glycolysis, and cholesterol biosynthesis pathways, suggesting that statins have pleiotropic effects on tumor cells. The above results suggest that repurposing statin drugs for ovarian cancer may provide a promising strategy to prevent and manage this devastating disease. ©2015 American Association for Cancer Research.

Engineering a functional 1-deoxy-D-xylulose 5-phosphate (DXP) pathway in Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Kirby, James; Dietzel, Kevin L.; Wichmann, Gale

2016-01-01

Isoprenoids are used in many commercial applications and much work has gone into engineering microbial hosts for their production. Isoprenoids are produced either from acetyl-CoA via the mevalonate pathway or from pyruvate and glyceraldehyde 3-phosphate via the 1-deoxy-D-xylulose 5-phosphate (DXP......) pathway. Saccharomyces cerevisiae exclusively utilizes the mevalonate pathway to synthesize native isoprenoids and in fact the alternative DXP pathway has never been found or successfully reconstructed in the eukaryotic cytosol. There are, however, several advantages to isoprenoid synthesis via the DXP...... time, functional expression of the DXP pathway in S. cerevisiae. Under low aeration conditions, an engineered strain relying solely on the DXP pathway for isoprenoid biosynthesis achieved an endpoint biomass 80% of that of the same strain using the mevalonate pathway....

Mevalonate Pathway Antagonist Inhibits Proliferation of Serous Tubal Intraepithelial Carcinoma and Ovarian Carcinoma in Mouse Models

Science.gov (United States)

Kobayashi, Yusuke; Kashima, Hiroyasu; Wu, Ren-Chin; Jung, Jin- Gyoung; Kuan, Jen-Chun; Gu, Jinghua; Xuan, Jianhua; Sokoll, Lori; Visvanathan, Kala; Shih, Ie-Ming; Wang, Tian-Li

2015-01-01

Purpose Statins are among the most frequently prescribed drugs because of their efficacy and low toxicity in treating hypercholesterolemia. Recently, statins have been reported to inhibit the proliferative activity of cancer cells, especially those with TP53 mutations. Since TP53 mutations occur in almost all of the ovarian high-grade serous carcinoma, we determined if statins suppressed tumor growth in animal models of ovarian cancer. Experimental Design Two ovarian cancer mouse models were employed. The first one was a genetically engineered model, mogp-TAg, in which the promoter of oviduct glycoprotein-1 was used to drive the expression of SV40 T-antigen in gynecologic tissues. These mice spontaneously develop serous tubal intraepithelial carcinomas (STICs), which are known as ovarian cancer precursor lesions. The second model was a xenograft tumor model in which human ovarian cancer cells were inoculated into immunocompromised mice. Mice in both models were treated with lovastatin, and effects on tumor growth were monitored. The molecular mechanisms underlying the anti-tumor effects of lovastatin were also investigated. Results Lovastatin significantly reduced the development of STICs in mogp-TAg mice and inhibited ovarian tumor growth in the mouse xenograft model. Knockdown of prenylation enzymes in the mevalonate pathway recapitulated the lovastatin-induced anti-proliferative phenotype. Transcriptome analysis indicated that lovastatin affected the expression of genes associated with DNA replication, Rho/PLC signaling, glycolysis, and cholesterol biosynthesis pathways, suggesting that statins have pleiotropic effects on tumor cells. Conclusion The above results suggest that repurposing statin drugs for ovarian cancer may provide a promising strategy to prevent and manage this devastating disease. PMID:26109099

Mevalonosomes: specific vacuoles containing the mevalonate pathway in Plocamium brasiliense cortical cells (Rhodophyta).

Science.gov (United States)

Paradas, Wladimir Costa; Crespo, Thalita Mendes; Salgado, Leonardo Tavares; de Andrade, Leonardo Rodrigues; Soares, Angélica Ribeiro; Hellio, Claire; Paranhos, Ricardo Rogers; Hill, Lilian Jorge; de Souza, Geysa Marinho; Kelecom, Alphonse Germaine Albert Charles; Da Gama, Bernardo Antônio Perez; Pereira, Renato Crespo; Amado-Filho, Gilberto Menezes

2015-04-01

This paper has identified, for the first time in a member of the Rhodophyta, a vacuolar organelle containing enzymes that are involved in the mevalonate pathway-an important step in red algal isoprenoid biosynthesis. These organelles were named mevalonosomes (Mev) and were found in the cortical cells (CC) of Plocamium brasiliense, a marine macroalgae that synthesizes several halogenated monoterpenes. P. brasiliense specimens were submitted to a cytochemical analysis of the activity of the 3-hydroxy-3-methylglutaryl-CoA synthase (HMGS). Using transmission electron microscopy (TEM), we confirmed the presence of HMGS activity within the Mev. Because HMGS is necessary for the biosynthesis of halogenated monoterpenes, we isolated a hexanic fraction (HF) rich in halogenated monoterpenes from P. brasiliense that contained a pentachlorinated monoterpene as a major metabolite. Because terpenes are often related to chemical defense, the antifouling (AF) activity of pentachlorinated monoterpene was tested. We found that the settlement of the mussel Perna perna was reduced by HF treatment (2.25 times less than control; 40% and 90% of fouled surface, respectively; P = 0.001; F9,9 = 1.13). The HF (at 10 μg · mL(-1) ) also inhibited three species of fouling microalgae (Chlorarachnion reptans, Cylindrotheca cloisterium, and Exanthemachrysis gayraliae), while at a higher concentration (50 μg · mL(-1) ), it inhibited the bacteria Halomonas marina, Polaribacter irgensii, Pseudoalteromonas elyakovii, Shewanella putrefaciens, and Vibrio aestuarianus. The AF activity of P. brasiliense halogenated monoterpenes and the localization of HMGS activity inside Mev suggest that this cellular structure found in CC may play a role in thallus protection against biofouling. © 2015 Phycological Society of America.

Propiconazole-enhanced hepatic cell proliferation is associated with dysregulation of the cholesterol biosynthesis pathway leading to activation of Erk1/2 through Ras farnesylation

Energy Technology Data Exchange (ETDEWEB)

Murphy, Lynea A.; Moore, Tanya; Nesnow, Stephen, E-mail: nesnow.stephen@epa.gov

2012-04-15

Propiconazole is a mouse hepatotumorigenic fungicide designed to inhibit CYP51, a key enzyme in the biosynthesis of ergosterol in fungi and is widely used in agriculture to prevent fungal growth. Metabolomic studies in mice revealed that propiconazole increased levels of hepatic cholesterol metabolites and bile acids, and transcriptomic studies revealed that genes within the cholesterol biosynthesis, cholesterol metabolism and bile acid biosyntheses pathways were up-regulated. Hepatic cell proliferation was also increased by propiconazole. AML12 immortalized hepatocytes were used to study propiconazole's effects on cell proliferation focusing on the dysregulation of cholesterol biosynthesis and resulting effects on Ras farnesylation and Erk1/2 activation as a primary pathway. Mevalonate, a key intermediate in the cholesterol biosynthesis pathway, increases cell proliferation in several cancer cell lines and tumors in vivo and serves as the precursor for isoprenoids (e.g. farnesyl pyrophosphate) which are crucial in the farnesylation of the Ras protein by farnesyl transferase. Farnesylation targets Ras to the cell membrane where it is involved in signal transduction, including the mitogen-activated protein kinase (MAPK) pathway. In our studies, mevalonic acid lactone (MVAL), a source of mevalonic acid, increased cell proliferation in AML12 cells which was reduced by farnesyl transferase inhibitors (L-744,832 or manumycin) or simvastatin, an HMG-CoA reductase inhibitor, indicating that this cell system responded to alterations in the cholesterol biosynthesis pathway. Cell proliferation in AML12 cells was increased by propiconazole which was reversed by co-incubation with L-744,832 or simvastatin. Increasing concentrations of exogenous cholesterol muted the proliferative effects of propiconazole and the inhibitory effects of L-733,832, results ascribed to reduced stimulation of the endogenous cholesterol biosynthesis pathway. Western blot analysis of subcellular

Liquid chromatography-tandem mass spectrometry method for the measurement of serum mevalonic acid: a novel marker of hydroxymethylglutaryl coenzyme A reductase inhibition by statins.

Science.gov (United States)

Waldron, Jenna; Webster, Craig

2011-05-01

Mevalonic acid (MVA) is synthesized at an early and rate-limiting step in the biosynthesis of cholesterol by the enzyme hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase, and is a useful measure of statin efficacy or treatment. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the measurement of serum MVA has been developed. Following the in vitro conversion of MVA to mevalonic acid lactone (MVAL) in the serum, MVAL and a deuterated internal standard were extracted using an online solid-phase extraction procedure. Chromatographic separation was achieved using a Luna PFP column (Phenomenex), with enhanced selectivity and improved resolution for polar compounds. A gradient system was used, with mobile phase comprising methanol and water (5 mmol/L ammonium formate buffer, pH 2.5). Analysis was performed using an API 5000 tandem mass spectrometer (Applied Biosystems) in positive electrospray ionization mode. The method showed excellent recoveries (98 ± 8%) and imprecision (intra-assay coefficient of variation of 2.2% [6.5 ng/mL] and 2.6% [10.5 ng/mL], and inter-assay coefficient of variation of 9% [10.5 ng/mL]). The assay provides a calibration range up to 50 ng/mL with a limit of detection at 0.1 ng/mL. A simple, rapid and analytically specific method has been developed for the measurement of serum MVA, in the form of MVAL. The high analytical sensitivity of the method allows for accurate quantitation of MVAL in serum samples, both at the endogenous levels found in healthy individuals and in statin-treated patients where normal levels are expected to be greatly reduced through the inhibition of HMG-CoA reductase.

Engineering a functional 1-deoxy-D-xylulose 5-phosphate (DXP) pathway in Saccharomyces cerevisiae

Energy Technology Data Exchange (ETDEWEB)

Kirby, James [Univ. of California, Berkeley, CA (United States). California Institute of Quantitative Biosciences (QB3); Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Dietzel, Kevin L. [Amyris, inc., Emeryville, CA (United States); Wichmann, Gale [Amyris, inc., Emeryville, CA (United States); Chan, Rossana [Univ. of California, Berkeley, CA (United States). California Institute of Quantitative Biosciences (QB3); Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Antipov, Eugene [Amyris, inc., Emeryville, CA (United States); Moss, Nathan [Amyris, inc., Emeryville, CA (United States); Baidoo, Edward E. K. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Jackson, Peter [Amyris, inc., Emeryville, CA (United States); Gaucher, Sara P. [Amyris, inc., Emeryville, CA (United States); Gottlieb, Shayin [Amyris, inc., Emeryville, CA (United States); LaBarge, Jeremy [Amyris, inc., Emeryville, CA (United States); Mahatdejkul, Tina [Amyris, inc., Emeryville, CA (United States); Hawkins, Kristy M. [Amyris, inc., Emeryville, CA (United States); Muley, Sheela [Amyris, inc., Emeryville, CA (United States); Newman, Jack D. [Amyris, inc., Emeryville, CA (United States); Liu, Pinghua [Boston Univ., MA (United States). Dept. of Chemistry; Keasling, Jay D. [Univ. of California, Berkeley, CA (United States). California Institute of Quantitative Biosciences (QB3); Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Univ. of California, Berkeley, CA (United States). Depts. of Chemical & Biomolecular Engineering and Bioengineering; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Biological Systems & Engineering Div.; Technical Univ. of Denmark, Hoesholm (Denmark). Novo Nodisk Foundation Center for Biosustainability; Zhao, Lishan [Amyris, inc., Emeryville, CA (United States)

2016-10-27

Isoprenoids are made by all free-living organisms and range from essential metabolites like sterols and quinones to more complex compounds like pinene and rubber. They are used in many commercial applications and much work has gone into engineering microbial hosts for their production. Isoprenoids are produced either from acetyl-CoA via the mevalonate pathway or from pyruvate and glyceraldehyde 3-phosphate via the 1-deoxy-D-xylulose 5-phosphate (DXP) pathway. Saccharomyces cerevisiae exclusively utilizes the mevalonate pathway to synthesize native isoprenoids and in fact the alternative DXP pathway has never been found or successfully reconstructed in the eukaryotic cytosol. There are, however, several advantages to isoprenoid synthesis via the DXP pathway, such as a higher theoretical yield, and it has long been a goal to transplant the pathway into yeast. In this work, we investigate and address barriers to DXP pathway functionality in S. cerevisiae using a combination of synthetic biology, biochemistry and metabolomics. We report, for the first time, functional expression of the DXP pathway in S. cerevisiae. Under low aeration conditions, an engineered strain relying solely on the DXP pathway for isoprenoid biosynthesis achieved an endpoint biomass 80% of that of the same strain using the mevalonate pathway.

Expression of the mevalonate pathway enzymes in the Lutzomyia longipalpis (Diptera: Psychodidae) sex pheromone gland demonstrated by an integrated proteomic approach.

Science.gov (United States)

González-Caballero, Natalia; Rodríguez-Vega, Andrés; Dias-Lopes, Geovane; Valenzuela, Jesus G; Ribeiro, Jose M C; Carvalho, Paulo Costa; Valente, Richard H; Brazil, Reginaldo P; Cuervo, Patricia

2014-01-16

In Latin America, Lutzomyia longipalpis is the main vector of the protozoan parasite Leishmania infantum, which is the causal agent of American Visceral Leishmaniasis. This insect uses male-produced pheromones for mate recognition. Elucidation of pheromone biogenesis or its regulation may enable molecular strategies for mating disruption and, consequently, the vector's population management. Motivated by our recent results of the transcriptomic characterization of the L. longipalpis pheromone gland, we performed a proteomic analysis of this tissue combining SDS-PAGE, and mass spectrometry followed by an integrative data analysis. Considering that annotated genome sequences of this sand fly are not available, we designed an alternative workflow searching MS/MS data against two customized databases using three search engines: Mascot, OMSSA and ProLuCID. A total of 542 proteins were confidently characterized, 445 of them using a Uniref100-insect protein database, and 97 using a transcript translated database. In addition, use of PEAKS for de novo peptide sequencing of MS/MS data confirmed ~90% identifications made with the combination of the three search engines. Our results include the identification of six of the seven enzymes of the mevalonate-pathway, plus the enzymes involved in sesquiterpenoid biosynthesis, all of which are proposed to be involved in pheromone production in L. longipalpis. L. longipalpis is the main vector of the protozoan parasite L. infantum, which is the causal agent of American Visceral Leishmaniasis. One of the control measures of such disease is focused on vector population control. As this insect uses male-produced pheromones for mate recognition, the elucidation of pheromone biogenesis or its regulating process may enable molecular strategies for mating disruption and, consequently, this vector's population management. On this regard, in this manuscript we report expression evidence, at the protein level, of several molecules potentially

Lack of Prenylated Proteins, Autophagy Impairment and Apoptosis in SH-SY5Y Neuronal Cell Model of Mevalonate Kinase Deficiency

Directory of Open Access Journals (Sweden)

Paola Maura Tricarico

2017-03-01

Full Text Available Background/Aims: Mevalonate Kinase Deficiency (MKD, is a hereditary disease due to mutations in mevalonate kinase gene (MVK. MKD has heterogeneous clinical phenotypes: the correlation between MVK mutations and MKD clinical phenotype is still to be fully elucidated. Deficiency of prenylated proteins has been hypothesized as possible MKD pathogenic mechanism. Based on this hypothesis and considering that neurologic impairment characterizes Mevalonic Aciduria (MA, the most severe form of MKD, we studied the effects of I268T and N301T MVK mutations on protein prenylation, autophagy and programmed cell death in SH-SY5Y neuroblastoma cell lines. Methods: SH-SY5Y cells were transiently transfected, with the pCMV-6 plasmid containing MVK wild type and the two mutated sequences. Protein prenylation levels were evaluated using GFP-RhoA-F to assess farnesylation, and GFP-RhoA to evaluate geranylgeranylation; autophagy was measured by evaluating LC3 and p62 protein levels, while Annexin V-FITC and Propidium Iodide staining allowed apoptosis detection. Results: MVK mutants’ over-expression causes decreased levels of farnesylation and geranylgeranylation, and also increased LC3 lipidation in SH-SY5Y, with concomitant p62 accumulation. Treatment with bafilomycin A1 (an inhibitor of vacuolar H+-ATPase, a late autophagy inhibitor further increase LC3-II and p62 levels, suggesting that degradation of autophagolysosome could be impaired. SH-SY5Y, with both MVK mutants, showed apoptosis increase; the presence of N301T associated with augmented cell death. Conclusions: We hypothesize that mevalonate pathway impairment causes alteration of farnesylation and geranylgeranylation proteins and alteration of the autophagic flux; these changes can induce apoptosis, possibly more relevant in the presence of N301T mutation.

Lack of Prenylated Proteins, Autophagy Impairment and Apoptosis in SH-SY5Y Neuronal Cell Model of Mevalonate Kinase Deficiency.

Science.gov (United States)

Tricarico, Paola Maura; Romeo, Alessandra; Gratton, Rossella; Crovella, Sergio; Celsi, Fulvio

2017-01-01

Mevalonate Kinase Deficiency (MKD), is a hereditary disease due to mutations in mevalonate kinase gene (MVK). MKD has heterogeneous clinical phenotypes: the correlation between MVK mutations and MKD clinical phenotype is still to be fully elucidated. Deficiency of prenylated proteins has been hypothesized as possible MKD pathogenic mechanism. Based on this hypothesis and considering that neurologic impairment characterizes Mevalonic Aciduria (MA), the most severe form of MKD, we studied the effects of I268T and N301T MVK mutations on protein prenylation, autophagy and programmed cell death in SH-SY5Y neuroblastoma cell lines. SH-SY5Y cells were transiently transfected, with the pCMV-6 plasmid containing MVK wild type and the two mutated sequences. Protein prenylation levels were evaluated using GFP-RhoA-F to assess farnesylation, and GFP-RhoA to evaluate geranylgeranylation; autophagy was measured by evaluating LC3 and p62 protein levels, while Annexin V-FITC and Propidium Iodide staining allowed apoptosis detection. MVK mutants' over-expression causes decreased levels of farnesylation and geranylgeranylation, and also increased LC3 lipidation in SH-SY5Y, with concomitant p62 accumulation. Treatment with bafilomycin A1 (an inhibitor of vacuolar H+-ATPase, a late autophagy inhibitor) further increase LC3-II and p62 levels, suggesting that degradation of autophagolysosome could be impaired. SH-SY5Y, with both MVK mutants, showed apoptosis increase; the presence of N301T associated with augmented cell death. We hypothesize that mevalonate pathway impairment causes alteration of farnesylation and geranylgeranylation proteins and alteration of the autophagic flux; these changes can induce apoptosis, possibly more relevant in the presence of N301T mutation. © 2017 The Author(s)Published by S. Karger AG, Basel.

Differential control of the cholesterol biosynthetic pathway in tumor versus liver: evidence for decontrolled tumor cholesterogenesis in a cell-free system

International Nuclear Information System (INIS)

Azrolan, N.

1987-01-01

Cholesterol biosynthesis was characterized in cell-free post-mitochondrial supernatant (PMS) systems prepared from both normal rat liver and Morris hepatoma 3924A. Per cell, the rate of cholesterol synthesis from either 14 C-citrate of 14 -acetate in the hepatoma system was 9-fold greater than that observed in the liver system. Furthermore, the ratio of sterol-to-fatty acid synthesis rates from 14 C-citrate was more than 3-fold greater in the tumor than in the normal liver system. Incubations using radiolabeled acetate and mevalonate have demonstrated the loss of a normally rate-limiting control site within the early portion of the cholesterol biosynthetic pathway in the tumor system. Upon analysis of the steady-state levels of early lipogenic intermediates, the specific site of decontrol in the tumor was identified as the 3-hydroxy-3-methylglutaryl-CoA → mevalonate site of this pathway. In contrast, this reaction appeared to retain its rate-limiting properties in the cell-free system from normal liver

Phylogenomic reconstruction of archaeal fatty acid metabolism

Science.gov (United States)

Dibrova, Daria V.; Galperin, Michael Y.; Mulkidjanian, Armen Y.

2014-01-01

While certain archaea appear to synthesize and/or metabolize fatty acids, the respective pathways still remain obscure. By analyzing the genomic distribution of the key lipid-related enzymes, we were able to identify the likely components of the archaeal pathway of fatty acid metabolism, namely, a combination of the enzymes of bacterial-type β-oxidation of fatty acids (acyl-CoA-dehydrogenase, enoyl-CoA hydratase, and 3-hydroxyacyl-CoA dehydrogenase) with paralogs of the archaeal acetyl-CoA C-acetyltransferase, an enzyme of the mevalonate biosynthesis pathway. These three β-oxidation enzymes working in the reverse direction could potentially catalyze biosynthesis of fatty acids, with paralogs of acetyl-CoA C-acetyltransferase performing addition of C2 fragments. The presence in archaea of the genes for energy-transducing membrane enzyme complexes, such as cytochrome bc complex, cytochrome c oxidase, and diverse rhodopsins, was found to correlate with the presence of the proposed system of fatty acid biosynthesis. We speculate that because these membrane complexes functionally depend on fatty acid chains, their genes could have been acquired via lateral gene transfer from bacteria only by those archaea that already possessed a system of fatty acid biosynthesis. The proposed pathway of archaeal fatty acid metabolism operates in extreme conditions and therefore might be of interest in the context of biofuel production and other industrial applications. PMID:24818264

Possible regulation of sterol biosynthesis by phenolic acids

International Nuclear Information System (INIS)

Ranganathan, S.; Ramasarma, T.

1974-01-01

To test whether the phenolic acids, metabolites of tyrosine, regulate the biosynthesis of cholesterol, influence of phenolic acids on the incorporation of mevalonate-2- 14 C into sterols by rat liver and brain homogenate systems has been investigated in vitro. Results show that the combined presence of the aromatic ring and the carboxyl group in the compound under investigation inhibited the incorporation of labelled mevalonate. (M.G.B.)

Analysis of iridoids content and expression studies of genes encoding early enzymes in the indol terpenoid biosynthesis pathway in Catharanthus roseus Análisis de iridoides y expresión de genes que codifican enzimas tempranas en la síntesis de alcaloides indol terpenoicos en Catharanthus roseus

OpenAIRE

Leech Mark; Palacios-Rojas Natalia

2004-01-01

Terpenoid indole alkaloids (TIA) are of pharmaceutical importance, however the industrial use of these compouds is very limited because its accumulation is very low in plant tissues. TIA are derived f rom the shikimate and terpenoid pathways, which supply secologanin and tryptamine, the indole and iridoid moieties, respectively. Secololganin is a terpenoid which is belived to be synthesised the MEP pathway rather than by the acetate/mevalonic acid pathway. Secologanin is thought to be a limit...

Crystal Structures of Staphylococcus epidermidis Mevalonate Diphosphate Decarboxylase Bound to Inhibitory Analogs Reveal New Insight into Substrate Binding and Catalysis

Energy Technology Data Exchange (ETDEWEB)

Barta, Michael L.; Skaff, D. Andrew; McWhorter, William J.; Herdendorf, Timothy J.; Miziorko, Henry M.; Geisbrecht, Brian V. (UMKC)

2011-10-28

The polyisoprenoid compound undecaprenyl phosphate is required for biosynthesis of cell wall peptidoglycans in Gram-positive bacteria, including pathogenic Enterococcus, Streptococcus, and Staphylococcus spp. In these organisms, the mevalonate pathway is used to produce the precursor isoprenoid, isopentenyl 5-diphosphate. Mevalonate diphosphate decarboxylase (MDD) catalyzes formation of isopentenyl 5-diphosphate in an ATP-dependent irreversible reaction and is therefore an attractive target for inhibitor development that could lead to new antimicrobial agents. To facilitate exploration of this possibility, we report the crystal structure of Staphylococcus epidermidis MDD (1.85 {angstrom} resolution) and, to the best of our knowledge, the first structures of liganded MDD. These structures include MDD bound to the mevalonate 5-diphosphate analogs diphosphoglycolyl proline (2.05 {angstrom} resolution) and 6-fluoromevalonate diphosphate (FMVAPP; 2.2 {angstrom} resolution). Comparison of these structures provides a physical basis for the significant differences in K{sub i} values observed for these inhibitors. Inspection of enzyme/inhibitor structures identified the side chain of invariant Ser{sup 192} as making potential contributions to catalysis. Significantly, Ser {yields} Ala substitution of this side chain decreases k{sub cat} by {approx}10{sup 3}-fold, even though binding interactions between FMVAPP and this mutant are similar to those observed with wild type MDD, as judged by the 2.1 {angstrom} cocrystal structure of S192A with FMVAPP. Comparison of microbial MDD structures with those of mammalian counterparts reveals potential targets at the active site periphery that may be exploited to selectively target the microbial enzymes. These studies provide a structural basis for previous observations regarding the MDD mechanism and inform future work toward rational inhibitor design.

Salicylic acid-independent plant defence pathways

OpenAIRE

Pieterse, C.M.J.; Loon, L.C. van

1999-01-01

Salicylic acid is an important signalling molecule involved in both locally and systemically induced disease resistance responses. Recent advances in our understanding of plant defence signalling have revealed that plants employ a network of signal transduction pathways, some of which are independent of salicylic acid. Evidence is emerging that jasmonic acid and ethylene play key roles in these salicylic acid-independent pathways. Cross-talk between the salicylic acid-dependent and the salicy...

Inhibition of Mevalonate Pathway and Synthesis of the Storage Lipids in Human Liver-Derived and Non-liver Cell Lines by Lippia alba Essential Oils.

Science.gov (United States)

Montero-Villegas, Sandra; Polo, Mónica; Galle, Marianela; Rodenak-Kladniew, Boris; Castro, María; Ves-Losada, Ana; Crespo, Rosana; García de Bravo, Margarita

2017-01-01

The essential oils (EOs) of Lippia alba, an herb extensively used as a folk medicine in Latin America, are today promoted as an effective means of eliminating problems caused by hyperlipemia. We hypothesized that L.alba EOs inhibited cholesterol and triacylglycerols synthesis and decreased the intracellular depots of those lipids (lipid droplets), mechanisms involving the induction of a hypolipidemic response. Our aim was, therefore, to evaluate the hypolipogenic capability of the EOs of four L. alba chemotypes on liver-derived (HepG2) and non-liver (A549) human cell lines and to identify the potential biochemical targets of those chemotypes, particularly within the mevalonate pathway (MP). [ 14 C]Acetate was used as radioactive precursor for assays. Lipid analyses were performed by thin-layer and capillary gas chromatography, lipid droplets analyzed by fluorescence microscopy, and HMGCR levels determined by Western blot. In both cell lines, all four chemotypes exerted hypocholesterogenic effects within a concentration range of 3.2-32 µg/mL. Nonsaponifiable lipids manifested a decrease in incorporation of [ 14 C]acetate into squalene, lanosterol, lathosterol, and cholesterol, but not into ubiquinone, thus suggesting an inhibition of enzymes in the MP downstream from farnesyl pyrophosphate. The tagetenone chemotype, the most efficacious hypocholesterogenic L. alba EO, lowered HMGCR protein levels; inhibited triacylglycerols, cholesteryl esters, and phospholipids synthesis; and diminished lipid droplets in size and volume. These results revealed that L. alba EOs inhibited different lipogenic pathways and such lipid-lowering effects could prove essential to prevent cardiovascular diseases.

Increasing production yield of tyrosine and mevalonate through inhibition of biomass formation

DEFF Research Database (Denmark)

Li, Songyuan; Jendresen, Christian Bille; Nielsen, Alex Toftgaard

2016-01-01

, in particular, resulted in an increase in mass yield of mevalonate and tyrosine by 80% and 50%, respectively. By tracking production and biomass concentrations, it was observed that the production was maintained for more than 10 h after inhibition of cell growth, despite cell maintenance requirements...

Identification of an itaconic acid degrading pathway in itaconic acid producing Aspergillus terreus.

Science.gov (United States)

Chen, Mei; Huang, Xuenian; Zhong, Chengwei; Li, Jianjun; Lu, Xuefeng

2016-09-01

Itaconic acid, one of the most promising and flexible bio-based chemicals, is mainly produced by Aspergillus terreus. Previous studies to improve itaconic acid production in A. terreus through metabolic engineering were mainly focused on its biosynthesis pathway, while the itaconic acid-degrading pathway has largely been ignored. In this study, we used transcriptomic, proteomic, bioinformatic, and in vitro enzymatic analyses to identify three key enzymes, itaconyl-CoA transferase (IctA), itaconyl-CoA hydratase (IchA), and citramalyl-CoA lyase (CclA), that are involved in the catabolic pathway of itaconic acid in A. terreus. In the itaconic acid catabolic pathway in A. terreus, itaconic acid is first converted by IctA into itaconyl-CoA with succinyl-CoA as the CoA donor, and then itaconyl-CoA is hydrated into citramalyl-CoA by IchA. Finally, citramalyl-CoA is cleaved into acetyl-CoA and pyruvate by CclA. Moreover, IctA can also catalyze the reaction between citramalyl-CoA and succinate to generate succinyl-CoA and citramalate. These results, for the first time, identify the three key enzymes, IctA, IchA, and CclA, involved in the itaconic acid degrading pathway in itaconic acid producing A. terreus. The results will facilitate the improvement of itaconic acid production by metabolically engineering the catabolic pathway of itaconic acid in A. terreus.

The ATP-binding cassette transporter A1 regulates phosphoantigen release and Vγ9Vδ2 T cell activation by dendritic cells

Science.gov (United States)

Castella, Barbara; Kopecka, Joanna; Sciancalepore, Patrizia; Mandili, Giorgia; Foglietta, Myriam; Mitro, Nico; Caruso, Donatella; Novelli, Francesco; Riganti, Chiara; Massaia, Massimo

2017-01-01

Vγ9Vδ2 T cells are activated by phosphoantigens, such as isopentenyl pyrophosphate (IPP), which is generated in the mevalonate pathway of antigen-presenting cells. IPP is released in the extracellular microenvironment via unknown mechanisms. Here we show that the ATP-binding cassette transporter A1 (ABCA1) mediates extracellular IPP release from dendritic cells (DC) in cooperation with apolipoprotein A-I (apoA-I) and butyrophilin-3A1. IPP concentrations in the supernatants are sufficient to induce Vγ9Vδ2 T cell proliferation after DC mevalonate pathway inhibition with zoledronic acid (ZA). ZA treatment increases ABCA1 and apoA-I expression via IPP-dependent LXRα nuclear translocation and PI3K/Akt/mTOR pathway inhibition. These results close the mechanistic gap in our understanding of extracellular IPP release from DC and provide a framework to fine-tune Vγ9Vδ2 T cell activation via mevalonate and PI3K/Akt/mTOR pathway modulation. PMID:28580927

Investigations on the isoprenoid biosynthesis in the green alga Scenedesmus obliquus by using the 13C-labelling technique

International Nuclear Information System (INIS)

Schwender, J.

1995-01-01

The biosynthesis of several prenyllipids (isoprenoid lipids) of the green alga Scendesmus obliquus was investigated. The aim was to verify, whether the biosynthesis of isopentenyl diphosphate (IPP) in Scenedesmus proceeds according to the classical acetate mevalonate pathway or to an alternative pathway. An alternative pathway for IPP formation has recently been detected in some eubacteria by the group of Prof. M. Rohmer. Some inhibition tests were performed with mevinolin, a specific inhibitor of HMG-CoA reductase which yields mevalonic acid. Mevinolin should block the biosynthesis of such isoprenoids which are formed via the acetate mevalonate pathway. Scenedesmus was grown heterotrophically on 13 C-labelled glucose or acetate. After isolation and purification of 13 C-labelled phytol (side chains of chlorophylls), β-carotene, lutein, plastoquinone-9 and three sterol compounds, the enrichment of 13 C at different carbon-positions of the labelled compounds was determined. This was achieved by the 13 C-NMR technique in cooperation with Miriam Seemann of the group of Prof. M. Rohmer in Mullhouse/France. (orig.) [de

Sterol-mediated regulation of mevalonic acid synthesis. Accumulation of 4-carboxysterols as the predominant sterols synthesized in a Chinese hamster ovary cell cholesterol auxotroph (mutant 215)

International Nuclear Information System (INIS)

Plemenitas, A.; Havel, C.M.; Watson, J.A.

1990-01-01

Chinese hamster ovary-215 (CHO-215) mutant cells are auxotrophic for cholesterol. Berry and Chang (Berry, D. J., and Chang, T. Y. (1982) Biochemistry 21, 573-580) suggested that the metabolic lesion was at the level of 4-methyl sterol oxidation. However, the observed cellular accumulation of lanosterol was not consistent with a defect at this metabolic site. With the use of a novel Silica Sep Pak sterol separation procedure, we demonstrated that 60-80% of the acetonesoluble lipid radioactivity in [5-3H]mevalonate-labeled CHO-215 cells was incorporated into acidic sterols. 7(8),Cholesten-4 beta-methyl,4 alpha-carboxy,3 beta-ol was the dominant end product. In addition to this acidic sterol, 7(8),24-cholestadien,4 beta-methyl,4 alpha-carboxy,3 beta-ol and 7(8),24-cholestadien,4 alpha-carboxy,3 beta-ol were also isolated. Incubation of cell-free extracts with [3H]7(8)-cholesten-4 beta-methyl, 4 alpha-carboxy,3 beta-ol and pyridine nucleotides confirmed that CHO-215 4-carboxysterol decarboxylase activity was less than 1% of that for wild type cells. Thus, a correspondence between decreased 4-carboxysterol decarboxylase activity and the spectrum of accumulated sterol products by intact CHO-215 cells was demonstrated. No detectable cholesterol was synthesized by CHO-215 cells. 3H-Product accumulation studies demonstrated that 7(8),24-cholestadien, 4 beta-methyl,4 alpha-carboxy,3 beta-ol increased prior to its subsequent saturation at the delta 24 carbon. Furthermore, the steady state ratio for delta 24-saturated acidic sterols/unsaturated acidic sterols was dependent on media cholesterol source and amount. Finally, the accumulated acidic sterol(s) were not regulatory signal molecules for the modulation of 3-hydroxy-3-methyl-glutaryl coenzyme. A reductase activity in response to cholesterol availability

Mevalonate-derived quinonemethide triterpenoid from in vitro roots of Peritassa laevigata and their localization in root tissue by MALDI imaging

Science.gov (United States)

Pina, Edieidia S.; Silva, Denise B.; Teixeira, Simone P.; Coppede, Juliana S.; Furlan, Maysa; França, Suzelei C.; Lopes, Norberto P.; Pereira, Ana Maria S.; Lopes, Adriana A.

2016-03-01

Biosynthetic investigation of quinonemethide triterpenoid 22β-hydroxy-maytenin (2) from in vitro root cultures of Peritassa laevigata (Celastraceae) was conducted using 13C-precursor. The mevalonate pathway in P. laevigata is responsible for the synthesis of the quinonemethide triterpenoid scaffold. Moreover, anatomical analysis of P. laevigata roots cultured in vitro and in situ showed the presence of 22β-hydroxy-maytenin (2) and maytenin (1) in the tissues from transverse or longitudinal sections with an intense orange color. MALDI-MS imaging confirmed the distribution of (2) and (1) in the more distal portions of the root cap, the outer cell layers, and near the vascular cylinder of P. laevigata in vitro roots suggesting a role in plant defense against infection by microorganisms as well as in the root exudation processes.

Skeletal muscle-specific HMG-CoA reductase knockout mice exhibit rhabdomyolysis: A model for statin-induced myopathy.

Science.gov (United States)

Osaki, Yoshinori; Nakagawa, Yoshimi; Miyahara, Shoko; Iwasaki, Hitoshi; Ishii, Akiko; Matsuzaka, Takashi; Kobayashi, Kazuto; Yatoh, Shigeru; Takahashi, Akimitsu; Yahagi, Naoya; Suzuki, Hiroaki; Sone, Hirohito; Ohashi, Ken; Ishibashi, Shun; Yamada, Nobuhiro; Shimano, Hitoshi

2015-10-23

HMG-CoA reductase (HMGCR) catalyzes the conversion of HMG-CoA to mevalonic acid (MVA); this is the rate-limiting enzyme of the mevalonate pathway that synthesizes cholesterol. Statins, HMGCR inhibitors, are widely used as cholesterol-reducing drugs. However, statin-induced myopathy is the most adverse side effect of statins. To eludicate the mechanisms underlying statin the myotoxicity and HMGCR function in the skeletal muscle, we developed the skeletal muscle-specific HMGCR knockout mice. Knockout mice exhibited postnatal myopathy with elevated serum creatine kinase levels and necrosis. Myopathy in knockout mice was completely rescued by the oral administration of MVA. These results suggest that skeletal muscle toxicity caused by statins is dependent on the deficiencies of HMGCR enzyme activity and downstream metabolites of the mevalonate pathway in skeletal muscles rather than the liver or other organs. Copyright © 2015 Elsevier Inc. All rights reserved.

Production of Odd-Carbon Dicarboxylic Acids in Escherichia coli Using an Engineered Biotin-Fatty Acid Biosynthetic Pathway.

Science.gov (United States)

Haushalter, Robert W; Phelan, Ryan M; Hoh, Kristina M; Su, Cindy; Wang, George; Baidoo, Edward E K; Keasling, Jay D

2017-04-05

Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids.

Integrating nitric oxide into salicylic acid and jasmonic acid/ethylene plant defense pathways

DEFF Research Database (Denmark)

Mur, Luis A J; Prats, Elena; Pierre, Sandra

2013-01-01

to be tailored to particular biotic stresses. Nitric oxide (NO) has emerged as a major signal influencing resistance mediated by both signalling pathways but no attempt has been made to integrate NO into established SA/JA/ET interactions. NO has been shown to act as an inducer or suppressor of signalling along......Plant defence against pests and pathogens is known to be conferred by either salicylic acid (SA) or jasmonic acid (JA)/ethylene (ET) pathways, depending on infection or herbivore-grazing strategy. It is well attested that SA and JA/ET pathways are mutually antagonistic allowing defence responses...

P53- and mevalonate pathway–driven malignancies require Arf6 for metastasis and drug resistance

Science.gov (United States)

Hashimoto, Ari; Oikawa, Tsukasa; Hashimoto, Shigeru; Sugino, Hirokazu; Yoshikawa, Ayumu; Otsuka, Yutaro; Handa, Haruka; Onodera, Yasuhito; Nam, Jin-Min; Oneyama, Chitose; Okada, Masato; Fukuda, Mitsunori

2016-01-01

Drug resistance, metastasis, and a mesenchymal transcriptional program are central features of aggressive breast tumors. The GTPase Arf6, often overexpressed in tumors, is critical to promote epithelial–mesenchymal transition and invasiveness. The metabolic mevalonate pathway (MVP) is associated with tumor invasiveness and known to prenylate proteins, but which prenylated proteins are critical for MVP-driven cancers is unknown. We show here that MVP requires the Arf6-dependent mesenchymal program. The MVP enzyme geranylgeranyl transferase II (GGT-II) and its substrate Rab11b are critical for Arf6 trafficking to the plasma membrane, where it is activated by receptor tyrosine kinases. Consistently, mutant p53, which is known to support tumorigenesis via MVP, promotes Arf6 activation via GGT-II and Rab11b. Inhibition of MVP and GGT-II blocked invasion and metastasis and reduced cancer cell resistance against chemotherapy agents, but only in cells overexpressing Arf6 and components of the mesenchymal program. Overexpression of Arf6 and mesenchymal proteins as well as enhanced MVP activity correlated with poor patient survival. These results provide insights into the molecular basis of MVP-driven malignancy. PMID:27044891

Salicylic acid-independent plant defence pathways

NARCIS (Netherlands)

Pieterse, C.M.J.; Loon, L.C. van

1999-01-01

Salicylic acid is an important signalling molecule involved in both locally and systemically induced disease resistance responses. Recent advances in our understanding of plant defence signalling have revealed that plants employ a network of signal transduction pathways, some of which are

Production of Odd-Carbon Dicarboxylic Acids in Escherichia coli Using an Engineered Biotin–Fatty Acid Biosynthetic Pathway

Energy Technology Data Exchange (ETDEWEB)

Haushalter, Robert W. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Phelan, Ryan M. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Hoh, Kristina M. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Su, Cindy [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Wang, George [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Baidoo, Edward E. K. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Keasling, Jay D. [Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division

2017-03-14

Dicarboxylic acids are commodity chemicals used in the production of plastics, polyesters, nylons, fragrances, and medications. Bio-based routes to dicarboxylic acids are gaining attention due to environmental concerns about petroleum-based production of these compounds. Some industrial applications require dicarboxylic acids with specific carbon chain lengths, including odd-carbon species. Biosynthetic pathways involving cytochrome P450-catalyzed oxidation of fatty acids in yeast and bacteria have been reported, but these systems produce almost exclusively even-carbon species. Here in this paper we report a novel pathway to odd-carbon dicarboxylic acids directly from glucose in Escherichia coli by employing an engineered pathway combining enzymes from biotin and fatty acid synthesis. Optimization of the pathway will lead to industrial strains for the production of valuable odd-carbon diacids.

An insight into structural and functional characteristics of 3-hydoxy 3-methyl glutarylCoA reductase from Ocimum species

Directory of Open Access Journals (Sweden)

Shilpi Bansal

2017-10-01

Full Text Available Secondary metabolites, the biological compounds secreted by plants as an aid to support their growth and development under stress conditions or as a part of their defense mechanism, now hold equal importance for mankind who employs it immensely for medication, flavorings, aroma, etc. Wide applicability of these compounds instigates one to understand the biosynthesis, structure and regulation of these bioactive molecules. Terpenoids form the largest group of secondary metabolites which comprise of a wide range of structurally and functionally distinct metabolites synthesized either via mevalonate pathway or non-mevalonate pathway. Targeting a key regulatory enzyme of this pathway, modulation of which would alter the carbon flux would be beneficial to enhance our knowledge about the above issue. For this the transcriptome (from SRA of different Ocimum species was mined out for important pathway genes using various bioinformatics approaches. Amongst them 3-hydoxy 3-methyl glutaryl CoA reductase (HMGR was selected which is the rate limiting enzyme in mevalonate pathway which controls the conversion of HMG-CoA to mevalonic acid. Isolation, cloning, protein expression, purification, etc. would be discussed in detail in the meeting. Full length protein was also characterized through bioinformatics tools to study its structure, properties, conserved domains, etc. Increase in secondary metabolite production by alteration of HMGR pool along with transcript modulation studies in planta revealed that HMGR gene governs the biosynthesis of secondary metabolites. Transcriptome mapping of different HMGR homologs which on comparison within member of same genus revealed its divergent nature which could account to its multifunctional role in different plants. Besides, providing a deep insight about the enzyme function combination of such molecular, transgenic and bioinformatics tools would help to develop strategies to engineer the HMGR mediated flux and also

Crosstalk between MAV and MEP pathways in vitro grape plants exposed to UV-B radiation

International Nuclear Information System (INIS)

Gil, M.; Bottini, R.; Piccoli, P.; Pontin, M.

2010-01-01

The synthesis of terpenoids from IPP (isopentenyl diphosphate) proceeds in plants throughout two pathways, the MVA (mevalonic acid) in cytosol and the MEP (2-C-methyl-D-erythritol 4-phosphate) in plastids. Ultraviolet-B (UV-B) radiation induced the synthesis of terpenes in in vitro grape plants according to the fluence rate. Low intensity UV-B promoted the MVA pathway while high intensity UV-B stimulated the MEP pathway. Mevastatin is known to inhibit the enzyme HMG-CoA reductase blocking terpene synthesis in cytosol. In vitro plants growing 45 days under 16 h-photoperiod (100 μmol m - 2 s - 1) were fed at the apex with mevastatin and then exposed to an UV-B dose administrated at two intensities: low UV-B (8.25 μW cm - 2,16 h) or high UV-B (33 μW cm - 2,4 h). Methanol: chloroform extracts were analyzed by GC-EIMS and compared with controls without mevastatin. Levels of γ-Sitosterol and Stigmasterol were significantly increased under low intensity UV-B in the controls. The plants treated with the inhibitor showed a significant decrease of both sterols and a decrease in the plastidial terpenes but sterols were higher under UV-B. These results suggest an IPP crosstalk between the MAV and MEP pathways under restrictive conditions. (authors)

Production of 3-Oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic Acid in the Fungus Aspergillus oryzae: A Step Towards Heterologous Production of Pyrethrins in Fungi.

Science.gov (United States)

Mohamed, Maged E; Pahirulzaman, Khomaizon A K; Lazarus, Colin M

2016-03-01

Pyrethrins are natural insecticides, which accumulate to high concentrations in pyrethrum (Chrysanthemum cinerariaefolium) flowers. Synthetic pyrethroids are more stable, more efficacious and cheaper, but contemporary requirements for safe and environmentally friendly pesticides encourage a return to the use of natural pyrethrins, and this would be favoured by development of an efficient route to their production by microbial fermentation. The biosynthesis of pyrethrins involves ester linkage between an acid moiety (chrysanthemoyl or pyrethroyl, synthesised via the mevalonic acid pathway from glucose), and an alcohol (pyrethrolone). Pyrethrolone is generated from 3-oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic acid, which originates from α-linolenic acid via the jasmonic acid biosynthetic cascade. The first four genes in this cascade, encoding lipoxygenase 2, allene-oxide synthase, allene-oxide cyclase 2 and 12-oxophytodienoic acid reductase 3, were amplified from an Arabidopsis thaliana cDNA library, cloned in a purpose-built fungal multigene expression vector and expressed in Aspergillus oryzae. HPLC-MS analysis of the transgenic fungus homogenate gave good evidence for the presence of 3-oxo-2-(2'-pentenyl)-cyclopentane-1-octanoic acid.

Cytochrome and Alternative Pathway Respiration in Tobacco (Effects of Salicylic Acid).

Science.gov (United States)

Rhoads, D. M.; McIntosh, L.

1993-11-01

In suspension cultures of NT1 tobacco (Nicotiana tabacum L. cv Bright Yellow) cells the cytochrome pathway capacity increased between d 3 and d 4 following subculturing and reached the highest level observed on d 7. The capacity decreased significantly by d 10 and was at the same level on d 14. Both alternative pathway capacity and the amount of the 35-kD alternative oxidase protein increased significantly between d 5 and d 6, reached the highest point observed on d 7, remained constant until d 10, and decreased by d 14. The highest capacities of the alternative and cytochrome pathways and the highest amount of the 35-kD protein were attained on the day that cell cultures reached a stationary phase of growth. Addition of salicylic acid to cell cultures on d 4 caused a significant increase in alternative pathway capacity and a dramatic accumulation of the 35-kD protein by 12 h. The alternative pathway capacity and the protein level reached the highest level observed by 16 h after salicylic acid addition, and the cytochrome pathway capacity was at about the same level at each time point. The accumulation of the 35-kD alternative oxidase protein was significantly decreased by addition of actinomycin D 1 h before salicylic acid and was blocked by addition of cycloheximide. These results indicate that de novo transcription and translation were necessary for salicylic acid to cause the maximum accumulation of the 35-kD protein.

Sterol homeostasis requires regulated degradation of squalene monooxygenase by the ubiquitin ligase Doa10/Teb4

Science.gov (United States)

Foresti, Ombretta; Ruggiano, Annamaria; Hannibal-Bach, Hans K; Ejsing, Christer S; Carvalho, Pedro

2013-01-01

Sterol homeostasis is essential for the function of cellular membranes and requires feedback inhibition of HMGR, a rate-limiting enzyme of the mevalonate pathway. As HMGR acts at the beginning of the pathway, its regulation affects the synthesis of sterols and of other essential mevalonate-derived metabolites, such as ubiquinone or dolichol. Here, we describe a novel, evolutionarily conserved feedback system operating at a sterol-specific step of the mevalonate pathway. This involves the sterol-dependent degradation of squalene monooxygenase mediated by the yeast Doa10 or mammalian Teb4, a ubiquitin ligase implicated in a branch of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway. Since the other branch of ERAD is required for HMGR regulation, our results reveal a fundamental role for ERAD in sterol homeostasis, with the two branches of this pathway acting together to control sterol biosynthesis at different levels and thereby allowing independent regulation of multiple products of the mevalonate pathway. DOI: http://dx.doi.org/10.7554/eLife.00953.001 PMID:23898401

Analyzing the structural aspects of Isoprenoid biosynthesis pathway proteins in Ocimum species

Directory of Open Access Journals (Sweden)

Muktesh Chandra

2017-10-01

Full Text Available Generally thought that the extremely diverse array of secondary metabolites observed within Ocimum species defends against a comparable diverse array of biotic pests, pathogens and herbivores encountered around its natural range. Along with defense the diverse array of secondary metabolite also leads to the therapeutic and remedial property which justifies Ocimum as natural medicinal and aromatic casket. Many of the defense compounds, aroma compounds and medicinal derivatives are secondary metabolites isolated from trichome glands, mainly consist of terpenoids as well as phenylpropanoids. Various pathways fabricating these compounds are known viz. mevalonate pathway (MVA, phenylpropanoid pathway and MEP pathways. The enzyme cascade responsible for various secondary metabolites, need to be explored in various aspects. Here we had studied the MVA pathway enzymes in O. basilicum and O. gratissimum to figure out variations in enzyme structures due to speciation. Hence, in depth analysis of the transcriptome of O. basilicum and O. gratissimum, varrying in qualitative and quantitative aspects of essential oil were carried out. The transcriptome data from NCBI server was assembled using bioinformatic approaches. nr database at NCBI repository used for annotation, which assigned 60% contigs to known functions. Contigs corresponding to Mevalonate pathway enzymes are isolated using perl pipelines developed in our lab, which were further assembled using CLC workbench to remove redundancy and make larger stretch of sequence. Blastx of these larger sequences assigned them function and they are mapped to validated sequences to make full length. Data from both species led us to overall seven enzymes (total 14 of MVA pathway. These enzymes are studied in detail for various physio-chemical properties, steriochemical properties and motif/domain for protein-protein interaction (PPI study. Homolog models of all enzymes were predicted, against templates from RCSB

Engineering the fatty acid metabolic pathway in Saccharomyces cerevisiae for advanced biofuel production

Directory of Open Access Journals (Sweden)

Xiaoling Tang

2015-12-01

Full Text Available Fatty acid-derived fuels and chemicals have attracted a great deal of attention in recent decades, due to their following properties of high compatibility to gasoline-based fuels and existing infrastructure for their direct utilization, storage and distribution. The yeast Saccharomyces cerevisiae is the ideal biofuel producing candidate, based on the wealth of available genetic information and versatile tools designed to manipulate its metabolic pathways. Engineering the fatty acid metabolic pathways in S. cerevisiae is an effective strategy to increase its fatty acid biosynthesis and provide more pathway precursors for production of targeted products. This review summarizes the recent progress in metabolic engineering of yeast cells for fatty acids and fatty acid derivatives production, including the regulation of acetyl-CoA biosynthesis, NADPH production, fatty acid elongation, and the accumulation of activated precursors of fatty acids for converting enzymes. By introducing specific enzymes in the engineered strains, a powerful platform with a scalable, controllable and economic route for advanced biofuel production has been established. Keywords: Metabolic engineering, Fatty acid biosynthesis, Fatty acid derivatives, Saccharomyces cerevisiae

Loss of HMG-CoA reductase in C. elegans causes defects in protein prenylation and muscle mitochondria.

Directory of Open Access Journals (Sweden)

Parmida Ranji

Full Text Available HMG-CoA reductase is the rate-limiting enzyme in the mevalonate pathway and the target of cholesterol-lowering statins. We characterized the C. elegans hmgr-1(tm4368 mutant, which lacks HMG-CoA reductase, and show that its phenotypes recapitulate that of statin treatment, though in a more severe form. Specifically, the hmgr-1(tm4368 mutant has defects in growth, reproduction and protein prenylation, is rescued by exogenous mevalonate, exhibits constitutive activation of the UPRer and requires less mevalonate to be healthy when the UPRmt is activated by a constitutively active form of ATFS-1. We also show that different amounts of mevalonate are required for different physiological processes, with reproduction requiring the highest levels. Finally, we provide evidence that the mevalonate pathway is required for the activation of the UPRmt.

Filling gaps in bacterial amino acid biosynthesis pathways with high-throughput genetics.

Directory of Open Access Journals (Sweden)

Morgan N Price

2018-01-01

Full Text Available For many bacteria with sequenced genomes, we do not understand how they synthesize some amino acids. This makes it challenging to reconstruct their metabolism, and has led to speculation that bacteria might be cross-feeding amino acids. We studied heterotrophic bacteria from 10 different genera that grow without added amino acids even though an automated tool predicts that the bacteria have gaps in their amino acid synthesis pathways. Across these bacteria, there were 11 gaps in their amino acid biosynthesis pathways that we could not fill using current knowledge. Using genome-wide mutant fitness data, we identified novel enzymes that fill 9 of the 11 gaps and hence explain the biosynthesis of methionine, threonine, serine, or histidine by bacteria from six genera. We also found that the sulfate-reducing bacterium Desulfovibrio vulgaris synthesizes homocysteine (which is a precursor to methionine by using DUF39, NIL/ferredoxin, and COG2122 proteins, and that homoserine is not an intermediate in this pathway. Our results suggest that most free-living bacteria can likely make all 20 amino acids and illustrate how high-throughput genetics can uncover previously-unknown amino acid biosynthesis genes.

Targeting arachidonic acid pathway by natural products for cancer prevention and therapy.

Science.gov (United States)

Yarla, Nagendra Sastry; Bishayee, Anupam; Sethi, Gautam; Reddanna, Pallu; Kalle, Arunasree M; Dhananjaya, Bhadrapura Lakkappa; Dowluru, Kaladhar S V G K; Chintala, Ramakrishna; Duddukuri, Govinda Rao

2016-10-01

Arachidonic acid (AA) pathway, a metabolic process, plays a key role in carcinogenesis. Hence, AA pathway metabolic enzymes phospholipase A 2 s (PLA 2 s), cyclooxygenases (COXs) and lipoxygenases (LOXs) and their metabolic products, such as prostaglandins and leukotrienes, have been considered novel preventive and therapeutic targets in cancer. Bioactive natural products are a good source for development of novel cancer preventive and therapeutic drugs, which have been widely used in clinical practice due to their safety profiles. AA pathway inhibitory natural products have been developed as chemopreventive and therapeutic agents against several cancers. Curcumin, resveratrol, apigenin, anthocyans, berberine, ellagic acid, eugenol, fisetin, ursolic acid, [6]-gingerol, guggulsteone, lycopene and genistein are well known cancer chemopreventive agents which act by targeting multiple pathways, including COX-2. Nordihydroguaiaretic acid and baicalein can be chemopreventive molecules against various cancers by inhibiting LOXs. Several PLA 2 s inhibitory natural products have been identified with chemopreventive and therapeutic potentials against various cancers. In this review, we critically discuss the possible utility of natural products as preventive and therapeutic agents against various oncologic diseases, including prostate, pancreatic, lung, skin, gastric, oral, blood, head and neck, colorectal, liver, cervical and breast cancers, by targeting AA pathway. Further, the current status of clinical studies evaluating AA pathway inhibitory natural products in cancer is reviewed. In addition, various emerging issues, including bioavailability, toxicity and explorability of combination therapy, for the development of AA pathway inhibitory natural products as chemopreventive and therapeutic agents against human malignancy are also discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Pimelic acid, the first precursor of the Bacillus subtilis biotin synthesis pathway, exists as the free acid and is assembled by fatty acid synthesis.

Science.gov (United States)

Manandhar, Miglena; Cronan, John E

2017-05-01

Biotin synthetic pathways are readily separated into two stages, synthesis of the seven carbon α, ω-dicarboxylic acid pimelate moiety and assembly of the fused heterocyclic rings. The biotin pathway genes responsible for pimelate moiety synthesis vary widely among bacteria whereas the ring synthesis genes are highly conserved. Bacillus subtilis seems to have redundant genes, bioI and bioW, for generation of the pimelate intermediate. Largely consistent with previous genetic studies it was found that deletion of bioW caused a biotin auxotrophic phenotype whereas deletion of bioI did not. BioW is a pimeloyl-CoA synthetase that converts pimelic acid to pimeloyl-CoA. The essentiality of BioW for biotin synthesis indicates that the free form of pimelic acid is an intermediate in biotin synthesis although this is not the case in E. coli. Since the origin of pimelic acid in Bacillus subtilis is unknown, 13 C-NMR studies were carried out to decipher the pathway for its generation. The data provided evidence for the role of free pimelate in biotin synthesis and the involvement of fatty acid synthesis in pimelate production. Cerulenin, an inhibitor of the key fatty acid elongation enzyme, FabF, markedly decreased biotin production by B. subtilis resting cells whereas a strain having a cerulenin-resistant FabF mutant produced more biotin. In addition, supplementation with pimelic acid fully restored biotin production in cerulenin-treated cells. These results indicate that pimelic acid originating from fatty acid synthesis pathway is a bona fide precursor of biotin in B. subtilis. © 2017 John Wiley & Sons Ltd.

Hexanoic acid is a resistance inducer that protects tomato plants against Pseudomonas syringae by priming the jasmonic acid and salicylic acid pathways.

Science.gov (United States)

Scalschi, Loredana; Vicedo, Begonya; Camañes, Gemma; Fernandez-Crespo, Emma; Lapeña, Leonor; González-Bosch, Carmen; García-Agustín, Pilar

2013-05-01

Hexanoic acid-induced resistance (Hx-IR) is effective against several pathogens in tomato plants. Our study of the mechanisms implicated in Hx-IR against Pseudomonas syringae pv. tomato DC3000 suggests that hexanoic acid (Hx) treatment counteracts the negative effect of coronatine (COR) and jasmonyl-isoleucine (JA-Ile) on the salicylic acid (SA) pathway. In Hx-treated plants, an increase in the expression of jasmonic acid carboxyl methyltransferase (JMT) and the SA marker genes PR1 and PR5 indicates a boost in this signalling pathway at the expense of a decrease in JA-Ile. Moreover, Hx treatment potentiates 12-oxo-phytodienoic acid accumulation, which suggests that this molecule might play a role per se in Hx-IR. These results support a positive relationship between the SA and JA pathways in Hx-primed plants. Furthermore, one of the mechanisms of virulence mediated by COR is stomatal re-opening on infection with P. syringae. In this work, we observed that Hx seems to inhibit stomatal opening in planta in the presence of COR, which suggests that, on infection in tomato, this treatment suppresses effector action to prevent bacterial entry into the mesophyll. © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

Analysis of Chlorogenic Acid Oxidation Pathway in Simulated ...

African Journals Online (AJOL)

Keywords: Honeysuckle, Chlorogenic acid, Enzymatic browning, Mimic system, Oxidation pathway, ... enzymatic oxidation of CA is the major cause of ..... to the concentration of catechol, o-quinone and current at PPO-modified microcylinder biosensor for diffusion- kinetic model. J Electroanal Chem 2011; 660: 200-208.

Sterol homeostasis requires regulated degradation of squalene monooxygenase by the ubiquitin ligase Doa10/Teb4

DEFF Research Database (Denmark)

Foresti, Ombretta; Ruggiano, Annamaria; Hannibal-Bach, Hans K

2013-01-01

Sterol homeostasis is essential for the function of cellular membranes and requires feedback inhibition of HMGR, a rate-limiting enzyme of the mevalonate pathway. As HMGR acts at the beginning of the pathway, its regulation affects the synthesis of sterols and of other essential mevalonate......-derived metabolites, such as ubiquinone or dolichol. Here, we describe a novel, evolutionarily conserved feedback system operating at a sterol-specific step of the mevalonate pathway. This involves the sterol-dependent degradation of squalene monooxygenase mediated by the yeast Doa10 or mammalian Teb4, a ubiquitin...... ligase implicated in a branch of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) pathway. Since the other branch of ERAD is required for HMGR regulation, our results reveal a fundamental role for ERAD in sterol homeostasis, with the two branches of this pathway acting together...

STEVIOSIDE SYNTHESIS IN STEVIA (Stevia rebaudiana Bert

Directory of Open Access Journals (Sweden)

Alfredo Jarma Orozco

2010-01-01

Full Text Available Stevia rebaudiana Bert. is a subtropical wild plant of Paraguay, that possesses a potent sweetener up to 300 times higher than sucrose and has no calories. The molecules responsible for these characteristics are diterpen glycosides, found in leaves and synthesized at least, at initial states, which use the same pathway of the gibberellic acid in which the shiquimic acid gives origin to many aromatic compounds. The acetate is the precursor of the terpens or isoprenoids through the acetate-mevalonate pathway, where steviosides are found. Although in this article, an alternative route is discussed. This updating pretends to contribute tools for the understanding of the main pathways of steviol glycosides synthesis.

Analysis of Chlorogenic Acid Oxidation Pathway in Simulated ...

African Journals Online (AJOL)

Purpose: To investigate the pathways involved in the oxidation of chlorogenic acid (CA) and phenol metabolism in honeysuckle buds. Methods: A model that mimics CA oxidation by honeysuckle polyphenol oxidase (PPO) by controlling the reaction temperature or reaction duration was employed, and the resulting products ...

Integrating nitric oxide into salicylic acid and jasmonic acid/ ethylene plant defense pathways.

Science.gov (United States)

Mur, Luis A J; Prats, Elena; Pierre, Sandra; Hall, Michael A; Hebelstrup, Kim H

2013-01-01

Plant defense against pests and pathogens is known to be conferred by either salicylic acid (SA) or jasmonic acid (JA)/ethylene (ET) pathways, depending on infection or herbivore-grazing strategy. It is well attested that SA and JA/ET pathways are mutually antagonistic allowing defense responses to be tailored to particular biotic stresses. Nitric oxide (NO) has emerged as a major signal influencing resistance mediated by both signaling pathways but no attempt has been made to integrate NO into established SA/JA/ET interactions. NO has been shown to act as an inducer or suppressor of signaling along each pathway. NO will initiate SA biosynthesis and nitrosylate key cysteines on TGA-class transcription factors to aid in the initiation of SA-dependent gene expression. Against this, S-nitrosylation of NONEXPRESSOR OF PATHOGENESIS-RELATED PROTEINS1 (NPR1) will promote the NPR1 oligomerization within the cytoplasm to reduce TGA activation. In JA biosynthesis, NO will initiate the expression of JA biosynthetic enzymes, presumably to over-come any antagonistic effects of SA on JA-mediated transcription. NO will also initiate the expression of ET biosynthetic genes but a suppressive role is also observed in the S-nitrosylation and inhibition of S-adenosylmethionine transferases which provides methyl groups for ET production. Based on these data a model for NO action is proposed but we have also highlighted the need to understand when and how inductive and suppressive steps are used.

Expression of Vibrio harveyi acyl-ACP synthetase allows efficient entry of exogenous fatty acids into the Escherichia coli fatty acid and lipid A synthetic pathways.

Science.gov (United States)

Jiang, Yanfang; Morgan-Kiss, Rachael M; Campbell, John W; Chan, Chi Ho; Cronan, John E

2010-02-02

Although the Escherichia coli fatty acid synthesis (FAS) pathway is the best studied type II fatty acid synthesis system, a major experimental limitation has been the inability to feed intermediates into the pathway in vivo because exogenously supplied free fatty acids are not efficiently converted to the acyl-acyl carrier protein (ACP) thioesters required by the pathway. We report that expression of Vibrio harveyi acyl-ACP synthetase (AasS), a soluble cytosolic enzyme that ligates free fatty acids to ACP to form acyl-ACPs, allows exogenous fatty acids to enter the E. coli fatty acid synthesis pathway. The free fatty acids are incorporated intact and can be elongated or directly incorporated into complex lipids by acyltransferases specific for acyl-ACPs. Moreover, expression of AasS strains and supplementation with the appropriate fatty acid restored growth to E. coli mutant strains that lack essential fatty acid synthesis enzymes. Thus, this strategy provides a new tool for circumventing the loss of enzymes essential for FAS function.

An Interferon Regulated MicroRNA Provides Broad Cell-Intrinsic Antiviral Immunity through Multihit Host-Directed Targeting of the Sterol Pathway.

Directory of Open Access Journals (Sweden)

Kevin A Robertson

2016-03-01

Full Text Available In invertebrates, small interfering RNAs are at the vanguard of cell-autonomous antiviral immunity. In contrast, antiviral mechanisms initiated by interferon (IFN signaling predominate in mammals. Whilst mammalian IFN-induced miRNA are known to inhibit specific viruses, it is not known whether host-directed microRNAs, downstream of IFN-signaling, have a role in mediating broad antiviral resistance. By performing an integrative, systematic, global analysis of RNA turnover utilizing 4-thiouridine labeling of newly transcribed RNA and pri/pre-miRNA in IFN-activated macrophages, we identify a new post-transcriptional viral defense mechanism mediated by miR-342-5p. On the basis of ChIP and site-directed promoter mutagenesis experiments, we find the synthesis of miR-342-5p is coupled to the antiviral IFN response via the IFN-induced transcription factor, IRF1. Strikingly, we find miR-342-5p targets mevalonate-sterol biosynthesis using a multihit mechanism suppressing the pathway at different functional levels: transcriptionally via SREBF2, post-transcriptionally via miR-33, and enzymatically via IDI1 and SC4MOL. Mass spectrometry-based lipidomics and enzymatic assays demonstrate the targeting mechanisms reduce intermediate sterol pathway metabolites and total cholesterol in macrophages. These results reveal a previously unrecognized mechanism by which IFN regulates the sterol pathway. The sterol pathway is known to be an integral part of the macrophage IFN antiviral response, and we show that miR-342-5p exerts broad antiviral effects against multiple, unrelated pathogenic viruses such Cytomegalovirus and Influenza A (H1N1. Metabolic rescue experiments confirm the specificity of these effects and demonstrate that unrelated viruses have differential mevalonate and sterol pathway requirements for their replication. This study, therefore, advances the general concept of broad antiviral defense through multihit targeting of a single host pathway.

An Interferon Regulated MicroRNA Provides Broad Cell-Intrinsic Antiviral Immunity through Multihit Host-Directed Targeting of the Sterol Pathway

Science.gov (United States)

Robertson, Kevin A.; Hsieh, Wei Yuan; Forster, Thorsten; Blanc, Mathieu; Lu, Hongjin; Crick, Peter J.; Yutuc, Eylan; Watterson, Steven; Martin, Kimberly; Griffiths, Samantha J.; Enright, Anton J.; Yamamoto, Mami; Pradeepa, Madapura M.; Lennox, Kimberly A.; Behlke, Mark A.; Talbot, Simon; Haas, Jürgen; Dölken, Lars; Griffiths, William J.; Wang, Yuqin; Angulo, Ana; Ghazal, Peter

2016-01-01

In invertebrates, small interfering RNAs are at the vanguard of cell-autonomous antiviral immunity. In contrast, antiviral mechanisms initiated by interferon (IFN) signaling predominate in mammals. Whilst mammalian IFN-induced miRNA are known to inhibit specific viruses, it is not known whether host-directed microRNAs, downstream of IFN-signaling, have a role in mediating broad antiviral resistance. By performing an integrative, systematic, global analysis of RNA turnover utilizing 4-thiouridine labeling of newly transcribed RNA and pri/pre-miRNA in IFN-activated macrophages, we identify a new post-transcriptional viral defense mechanism mediated by miR-342-5p. On the basis of ChIP and site-directed promoter mutagenesis experiments, we find the synthesis of miR-342-5p is coupled to the antiviral IFN response via the IFN-induced transcription factor, IRF1. Strikingly, we find miR-342-5p targets mevalonate-sterol biosynthesis using a multihit mechanism suppressing the pathway at different functional levels: transcriptionally via SREBF2, post-transcriptionally via miR-33, and enzymatically via IDI1 and SC4MOL. Mass spectrometry-based lipidomics and enzymatic assays demonstrate the targeting mechanisms reduce intermediate sterol pathway metabolites and total cholesterol in macrophages. These results reveal a previously unrecognized mechanism by which IFN regulates the sterol pathway. The sterol pathway is known to be an integral part of the macrophage IFN antiviral response, and we show that miR-342-5p exerts broad antiviral effects against multiple, unrelated pathogenic viruses such Cytomegalovirus and Influenza A (H1N1). Metabolic rescue experiments confirm the specificity of these effects and demonstrate that unrelated viruses have differential mevalonate and sterol pathway requirements for their replication. This study, therefore, advances the general concept of broad antiviral defense through multihit targeting of a single host pathway. PMID:26938778

Mechanisms of amino acid sensing in mTOR signaling pathway

OpenAIRE

Kim, Eunjung

2009-01-01

Amino acids are fundamental nutrients for protein synthesis and cell growth (increase in cell size). Recently, many compelling evidences have shown that the level of amino acids is sensed by extra- or intra-cellular amino acids sensor(s) and regulates protein synthesis/degradation. Mammalian target of rapamycin complex 1 (mTORC1) is placed in a central position in cell growth regulation and dysregulation of mTOR signaling pathway has been implicated in many serious human diseases including ca...

Viral MicroRNAs Repress the Cholesterol Pathway, and 25-Hydroxycholesterol Inhibits Infection.

Science.gov (United States)

Serquiña, Anna K P; Kambach, Diane M; Sarker, Ontara; Ziegelbauer, Joseph M

2017-07-11

From various screens, we found that Kaposi's sarcoma-associated herpesvirus (KSHV) viral microRNAs (miRNAs) target several enzymes in the mevalonate/cholesterol pathway. 3-Hydroxy-3-methylglutaryl-coenzyme A (CoA) synthase 1 (HMGCS1), 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR [a rate-limiting step in the mevalonate pathway]), and farnesyl-diphosphate farnesyltransferase 1 (FDFT1 [a committed step in the cholesterol branch]) are repressed by multiple KSHV miRNAs. Transfection of viral miRNA mimics in primary endothelial cells (human umbilical vein endothelial cells [HUVECs]) is sufficient to reduce intracellular cholesterol levels; however, small interfering RNAs (siRNAs) targeting only HMGCS1 did not reduce cholesterol levels. This suggests that multiple targets are needed to perturb this tightly regulated pathway. We also report here that cholesterol levels were decreased in de novo -infected HUVECs after 7 days. This reduction is at least partially due to viral miRNAs, since the mutant form of KSHV lacking 10 of the 12 miRNA genes had increased cholesterol compared to wild-type infections. We hypothesized that KSHV is downregulating cholesterol to suppress the antiviral response by a modified form of cholesterol, 25-hydroxycholesterol (25HC). We found that the cholesterol 25-hydroxylase (CH25H) gene, which is responsible for generating 25HC, had increased expression in de novo -infected HUVECs but was strongly suppressed in long-term latently infected cell lines. We found that 25HC inhibits KSHV infection when added exogenously prior to de novo infection. In conclusion, we found that multiple KSHV viral miRNAs target enzymes in the mevalonate pathway to modulate cholesterol in infected cells during latency. This repression of cholesterol levels could potentially be beneficial to viral infection by decreasing the levels of 25HC. IMPORTANCE A subset of viruses express unique microRNAs (miRNAs), which act like cellular miRNAs to generally repress host gene

Molecular cloning of a novel glucuronokinase/putative pyrophosphorylase from zebrafish acting in an UDP-glucuronic acid salvage pathway.

Directory of Open Access Journals (Sweden)

Roman Gangl

Full Text Available In animals, the main precursor for glycosaminoglycan and furthermore proteoglycan biosynthesis, like hyaluronic acid, is UDP-glucuronic acid, which is synthesized via the nucleotide sugar oxidation pathway. Mutations in this pathway cause severe developmental defects (deficiency in the initiation of heart valve formation. In plants, UDP-glucuronic acid is synthesized via two independent pathways. Beside the nucleotide sugar oxidation pathway, a second minor route to UDP-glucuronic acid exist termed the myo-inositol oxygenation pathway. Within this myo-inositol is ring cleaved into glucuronic acid, which is subsequently converted to UDP-glucuronic acid by glucuronokinase and UDP-sugar pyrophosphorylase. Here we report on a similar, but bifunctional enzyme from zebrafish (Danio rerio which has glucuronokinase/putative pyrophosphorylase activity. The enzyme can convert glucuronic acid into UDP-glucuronic acid, required for completion of the alternative pathway to UDP-glucuronic acid via myo-inositol and thus establishes a so far unknown second route to UDP-glucuronic acid in animals. Glucuronokinase from zebrafish is a member of the GHMP-kinase superfamily having unique substrate specificity for glucuronic acid with a Km of 31 ± 8 µM and accepting ATP as the only phosphate donor (Km: 59 ± 9 µM. UDP-glucuronic acid pyrophosphorylase from zebrafish has homology to bacterial nucleotidyltransferases and requires UTP as nucleosid diphosphate donor. Genes for bifunctional glucuronokinase and putative UDP-glucuronic acid pyrophosphorylase are conserved among some groups of lower animals, including fishes, frogs, tunicates, and polychaeta, but are absent from mammals. The existence of a second pathway for UDP-glucuronic acid biosynthesis in zebrafish likely explains some previous contradictory finding in jekyll/ugdh zebrafish developmental mutants, which showed residual glycosaminoglycans and proteoglycans in knockout mutants of UDP

Identification and preliminary characterization of protein-cysteine farnesyltransferase

International Nuclear Information System (INIS)

Manne, V.; Roberts, D.; Tobin, A.; O'Rourke, E.; Barbacid, M.; De Virgilio, M.; Meyers, C.; Ahmed, N.; Kurz, B.; Resh, M.; Kung, Hsiang-Fu

1990-01-01

Ras proteins must be isoprenylated at a conserved cysteine residue near the carboxyl terminus in order to exert their biological activity. Previous studies indicate that an intermediate in the mevalonate pathway, most likely farnesyl pyrophosphate, is the donor of this isoprenyl group. Inhibition of mevalonate synthesis reverts the abnormal phenotypes induced by the mutant RAS2 Valendash19 gene in Saccharomyces cerevisiae and blocks the maturation of Xenopus oocytes induced by an onocogenic Ras p21 protein of human origin. These results have raised the possibility of using inhibitors of the mevalonate pathway to block the transforming properties of ras oncogenes. Unfortunately, mevalonate is a precursor of various end products essential to mammalian cells, such as dolichols, ubiquinones, heme A, and cholesterol. In this study, the authors describe an enzymatic activity(ies) capable of catalyzing the farnesylation of unprocessed Ras p21 proteins in vitro at the correct (Cys-186) residue. Gel filtration analysis of a partially purified preparation of protein farnesyltransferase revealed two peaks of activity at 250-350 kDa and 80-130 kDa. Availability of an in vitro protein farnesyltransferase assay should be useful in screening for potential inhibitors of ras oncogene function that will not interfere with other aspects of the mevalonate pathway

Aluminum inhibits phosphatidic acid formation by blocking the phospholipase C pathway

NARCIS (Netherlands)

Ramos-Díaz, A.; Brito-Argáez, L.; Munnik, T.; Hernández-Sotomayor, S.M.T.

2007-01-01

Aluminum (Al(3+)) has been recognized as a main toxic factor in crop production in acid lands. Phosphatidic acid (PA) is emerging as an important lipid signaling molecule and has been implicated in various stress-signaling pathways in plants. In this paper, we focus on how PA generation is affected

Amino acids as regulators and components of nonproteinogenic pathways

NARCIS (Netherlands)

Meijer, Alfred J.

2003-01-01

Amino acids are not only important precursors for the synthesis of proteins and other N-containing compounds, but also participate in the regulation of major metabolic pathways. Glutamate and aspartate, for example, are components of the malate/aspartate shuttle and their concentrations control the

Metabolomics Analysis of the Toxic Effects of the Production of Lycopene and Its Precursors

Directory of Open Access Journals (Sweden)

April M. Miguez

2018-05-01

Full Text Available Using cells as microbial factories enables highly specific production of chemicals with many advantages over chemical syntheses. A number of exciting new applications of this approach are in the area of precision metabolic engineering, which focuses on improving the specificity of target production. In recent work, we have used precision metabolic engineering to design lycopene-producing Escherichia coli for use as a low-cost diagnostic biosensor. To increase precursor availability and thus the rate of lycopene production, we heterologously expressed the mevalonate pathway. We found that simultaneous induction of these pathways increases lycopene production, but induction of the mevalonate pathway before induction of the lycopene pathway decreases both lycopene production and growth rate. Here, we aim to characterize the metabolic changes the cells may be undergoing during expression of either or both of these heterologous pathways. After establishing an improved method for quenching E. coli for metabolomics analysis, we used two-dimensional gas chromatography coupled to mass spectrometry (GCxGC-MS to characterize the metabolomic profile of our lycopene-producing strains in growth conditions characteristic of our biosensor application. We found that the metabolic impacts of producing low, non-toxic levels of lycopene are of much smaller magnitude than the typical metabolic changes inherent to batch growth. We then used metabolomics to study differences in metabolism caused by the time of mevalonate pathway induction and the presence of the lycopene biosynthesis genes. We found that overnight induction of the mevalonate pathway was toxic to cells, but that the cells could recover if the lycopene pathway was not also heterologously expressed. The two pathways appeared to have an antagonistic metabolic effect that was clearly reflected in the cells’ metabolic profiles. The metabolites homocysteine and homoserine exhibited particularly interesting

The role of uric acid in the pathogenesis of diabetic retinopathy based on notch pathway.

Science.gov (United States)

Zhu, Dan-Dan; Wang, Yun-Zhi; Zou, Chen; She, Xin-Ping; Zheng, Zhi

2018-06-19

Uric acid has been proposed as an independent risk factor of diabetic retinopathy. Although Notch signaling was reported to be affected in the presence of high concentrations of uric acid or glucose, the underlying mechanisms of hyperuricemia through the Notch signaling pathway to promote the development of diabetic retinopathy remain unknown. We incubated human retinal endothelial cells (HRECs) with high glucose, high uric acid and high glucose plus high glucose respectively and evaluated the apoptosis rate in different treated cells by Tunel staining. We induced diabetic model by intraperitoneally streptozotocin. Then healthy rats and diabetic rats were given with adenine and oteracil potassium by gavage. Using automatic biochemical analyzer to detect blood glucose, uric acid, urea nitrogen, creatinine levels, to verify the success of modeling. The expression and mRNA levels of ICAM-1, IL-6, MCP-1, TNF-a, receptors Notch 1, ligands Dll 1, Dll 4, Jagged 1, Jagged 2 were detected by RT-PCR and Western-Blot. Notch1 siRNA was used to interfere Notch signaling pathway, the expression and mRNA levels of ICAM-1, IL-6, MCP-1 and TNF-α was detected by RT-PCR and Western blot respectively. In vitro models, the apoptosis of HRECs cells in high uric acid plus high glucose group was the most significant. In vitro and vivo models, detection of inflammatory cytokines revealed that the expression of inflammatory cytokines increased most significantly in high uric acid plus high glucose group. Notch signaling pathway activity was also increased most significantly in high uric acid plus high glucose group. After Notch 1 siRNA transfection in high glucose and high glucose plus uric acid group, the activity of Notch signaling pathway was successfully down-regulated. We found that the apoptosis of HRECs was significantly decreased in cells transfected with Notch 1 siRNA compared to the blank vector group, and the expression of inflammatory cytokines in cells was also significantly

Abscisic-acid-induced cellular apoptosis and differentiation in glioma via the retinoid acid signaling pathway.

Science.gov (United States)

Zhou, Nan; Yao, Yu; Ye, Hongxing; Zhu, Wei; Chen, Liang; Mao, Ying

2016-04-15

Retinoid acid (RA) plays critical roles in regulating differentiation and apoptosis in a variety of cancer cells. Abscisic acid (ABA) and RA are direct derivatives of carotenoids and share structural similarities. Here we proposed that ABA may also play a role in cellular differentiation and apoptosis by sharing a similar signaling pathway with RA that may be involved in glioma pathogenesis. We reported for the first time that the ABA levels were twofold higher in low-grade gliomas compared with high-grade gliomas. In glioma tissues, there was a positive correlation between the ABA levels and the transcription of cellular retinoic acid-binding protein 2 (CRABP2) and a negative correlation between the ABA levels and transcription of fatty acid-binding protein 5 (FABP5). ABA treatment induced a significant increase in the expression of CRABP2 and a decrease in the expression of peroxisome proliferator-activated receptor (PPAR) in glioblastoma cells. Remarkably, both cellular apoptosis and differentiation were increased in the glioblastoma cells after ABA treatment. ABA-induced cellular apoptosis and differentiation were significantly reduced by selectively silencing RAR-α, while RAR-α overexpression exaggerated the ABA-induced effects. These results suggest that ABA may play a role in the pathogenesis of glioma by promoting cellular apoptosis and differentiation through the RA signaling pathway. © 2015 UICC.

Nitrogen-containing bisphosphonate, YM529/ONO-5920 (a novel minodronic acid), inhibits RANKL expression in a cultured bone marrow stromal cell line ST2

International Nuclear Information System (INIS)

Nishida, Shozo; Tsubaki, Masanobu; Hoshino, Mayumi; Namimatsu, Ayumi; Uji, Hiromi; Yoshioka, Shohei; Tanimori, Yoshihiro; Yanae, Masashi; Iwaki, Masahiro; Irimajiri, Kiyohiro

2005-01-01

Increase in bone resorption by osteoclasts can cause metabolic bone diseases, such as osteoporosis. Recent attention has been paid to the receptor activator of the NF-κB ligand (RANKL), an accelerator of osteoclast differentiation. RANKL is expressed on the bone marrow-derived stromal cell membrane and induces the differentiation of osteoclasts by binding to RANK expressed on the osteoclast precursor cell membrane. Since the inhibition of RANKL expression can lead to the inhibition of osteoclastic bone resorption, the clinical application of RANKL inhibition could be expected to have a major effect on metabolic bone disease therapy. In this study, we investigated whether or not YM529/ONO-5920, a nitrogen-containing bisphosphonate (a novel minodronic acid), inhibits RANKL expression in a bone marrow-derived stromal cell line (ST2 cells). Reverse transcription-polymerase chain reaction revealed that the administration of YM529/ONO-5920 to ST2 cells inhibited RANKL mRNA expression and reduced RANKL proteins as assessed by Western blot analysis. The inhibition of RANKL mRNA expression was reversed when geranylgeranyl pyrophosphate (GGPP), an intermediate in the mevalonate pathway, was used in combination. Furthermore, YM529/ONO-5920 reduced phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), and similarly, U0126, a mitogen-activated protein kinase kinase 1/2 inhibitor, inhibited RANKL expression. Pretreatment with GGPP reversed the YM529/ONO-5920-induced decrease in phosphorylation of ERK. Furthermore, YM529/ONO-5920 decreased TRAP-positive cells in co-culture of ST2 cells and an osteoclast cell line, C7 cells, and this decrease was inhibited by pretreatment with GGPP. This indicates that YM529/ONO-5920 inhibits GGPP biosynthesis in the mevalonate pathway and then signal transduction in the Ras-mitogen-activated protein kinase pathway, thereby inhibiting RANKL expression on ST2 cells. These results suggest a newly elucidated action of bisphosphonates in

Microbial Community Pathways for the Production of Volatile Fatty Acids From CO2 and Electricity

Directory of Open Access Journals (Sweden)

Jorge Wenzel

2018-04-01

Full Text Available This study aims at elucidating the metabolic pathways involved in the production of volatile fatty acids from CO2 and electricity. Two bioelectrochemical systems (BES were fed with pure CO2 (cells A and B. The cathode potential was first poised at −574 mV vs. standard hydrogen electrode (SHE and then at −756 mV vs. SHE in order to ensure the required reducing power. Despite applying similar operation conditions to both BES, they responded differently. A mixture of organic compounds (1.87 mM acetic acid, 2.30 mM formic acid, 0.43 mM propionic acid, 0.15 mM butyric acid, 0.55 mM valeric acid, and 0.62 mM ethanol was produced in cell A while mainly 1.82 mM acetic acid and 0.23 mM propionic acid were produced in cell B. The microbial community analysis performed by 16S rRNA gene pyrosequencing showed a predominance of Clostridium sp. and Serratia sp. in cell A whereas Burkholderia sp. and Xanthobacter sp. predominated in cell B. The coexistence of three metabolic pathways involved in carbon fixation was predicted. Calvin cycle was predicted in both cells during the whole experiment while Wood-Ljungdahl and Arnon-Buchanan pathways predominated in the period with higher coulombic efficiency. Metabolic pathways which transform organic acids into anabolic intermediaries were also predicted, indicating the occurrence of complex trophic interactions. These results further complicate the understanding of these mixed culture microbial processes but also expand the expectation of compounds that could potentially be produced with this technology.

AMPK-independent pathways regulate skeletal muscle fatty acid oxidation

DEFF Research Database (Denmark)

Dzamko, Nicolas; Schertzer, Jonathan D.; Ryall, James G.

2008-01-01

The activation of AMP-activated protein kinase (AMPK) and phosphorylation/inhibition of acetyl-CoA carboxylase 2 (ACC2) is believed to be the principal pathway regulating fatty acid oxidation. However, during exercise AMPK activity and ACC Ser-221 phosphorylation does not always correlate...... with rates of fatty acid oxidation. To address this issue we have investigated the requirement for skeletal muscle AMPK in controlling aminoimidazole-4-carboxymide-1-beta-d-ribofuranoside (AICAR) and contraction-stimulated fatty acid oxidation utilizing transgenic mice expressing a muscle-specific kinase...... dead (KD) AMPK alpha2. In wild-type (WT) mice, AICAR and contraction increased AMPK alpha2 and alpha1 activities, the phosphorylation of ACC2 and rates of fatty acid oxidation while tending to reduce malonyl-CoA levels. Despite no activation of AMPK in KD mice, ACC2 phosphorylation was maintained...

Caffeic Acid Induces Apoptosis in Human Cervical Cancer Cells Through the Mitochondrial Pathway

Directory of Open Access Journals (Sweden)

Wei-Chun Chang

2010-12-01

Conclusion: Caffeic acid induces apoptosis by inhibiting Bcl-2 activity, leading to release of cytochrome c and subsequent activation of caspase-3, indicating that caffeic acid induces apoptosis via the mitochondrial apoptotic pathway. This also suggests that caffeic acid has a strong anti-tumor effect and may be a promising chemopreventive or chemotherapeutic agent.

Chitosan oligosaccharide induces resistance to Tobacco mosaic virus in Arabidopsis via the salicylic acid-mediated signalling pathway.

Science.gov (United States)

Jia, Xiaochen; Meng, Qingshan; Zeng, Haihong; Wang, Wenxia; Yin, Heng

2016-05-18

Chitosan is one of the most abundant carbohydrate biopolymers in the world, and chitosan oligosaccharide (COS), which is prepared from chitosan, is a plant immunity regulator. The present study aimed to validate the effect of COS on inducing resistance to tobacco mosaic virus (TMV) in Arabidopsis and to investigate the potential defence-related signalling pathways involved. Optimal conditions for the induction of TMV resistance in Arabidopsis were COS pretreatment at 50 mg/L for 1 day prior to inoculation with TMV. Multilevel indices, including phenotype data, and TMV coat protein expression, revealed that COS induced TMV resistance in wild-type and jasmonic acid pathway- deficient (jar1) Arabidopsis plants, but not in salicylic acid pathway deficient (NahG) Arabidopsis plants. Quantitative-PCR and analysis of phytohormone levels confirmed that COS pretreatment enhanced the expression of the defence-related gene PR1, which is a marker of salicylic acid signalling pathway, and increased the amount of salicylic acid in WT and jar1, but not in NahG plants. Taken together, these results confirm that COS induces TMV resistance in Arabidopsis via activation of the salicylic acid signalling pathway.

A nitrous acid biosynthetic pathway for diazo group formation in bacteria.

Science.gov (United States)

Sugai, Yoshinori; Katsuyama, Yohei; Ohnishi, Yasuo

2016-02-01

Although some diazo compounds have bioactivities of medicinal interest, little is known about diazo group formation in nature. Here we describe an unprecedented nitrous acid biosynthetic pathway responsible for the formation of a diazo group in the biosynthesis of the ortho-diazoquinone secondary metabolite cremeomycin in Streptomyces cremeus. This finding provides important insights into the biosynthetic pathways not only for diazo compounds but also for other naturally occurring compounds containing nitrogen-nitrogen bonds.

Permanganate oxidation of α-amino acids: kinetic correlations for the nonautocatalytic and autocatalytic reaction pathways.

Science.gov (United States)

Perez-Benito, Joaquin F

2011-09-08

The reactions of permanganate ion with seven α-amino acids in aqueous KH(2)PO(4)/K(2)HPO(4) buffers have been followed spectrophotometrically at two different wavelengths: 526 nm (decay of MnO(4)(-)) and 418 nm (formation of colloidal MnO(2)). All of the reactions studied were autocatalyzed by colloidal MnO(2), with the contribution of the autocatalytic reaction pathway decreasing in the order glycine > l-threonine > l-alanine > l-glutamic acid > l-leucine > l-isoleucine > l-valine. The rate constants corresponding to the nonautocatalytic and autocatalytic pathways were obtained by means of either a differential rate law or an integrated one, the latter requiring the use of an iterative method for its implementation. The activation parameters for the two pathways were determined and analyzed to obtain statistically significant correlations for the series of reactions studied. The activation enthalpy of the nonautocatalytic pathway showed a strong, positive dependence on the standard Gibbs energy for the dissociation of the protonated amino group of the α-amino acid. Linear enthalpy-entropy correlations were found for both pathways, leading to isokinetic temperatures of 370 ± 21 K (nonautocatalytic) and 364 ± 28 K (autocatalytic). Mechanisms in agreement with the experimental data are proposed for the two reaction pathways.

Saturated fatty acids enhance TLR4 immune pathways in human trophoblasts.

Science.gov (United States)

Yang, Xiaohua; Haghiac, Maricela; Glazebrook, Patricia; Minium, Judi; Catalano, Patrick M; Hauguel-de Mouzon, Sylvie

2015-09-01

What are the effects of fatty acids on placental inflammatory cytokine with respect to toll-like receptor-4/nuclear factor-kappa B (TLR4/NF-kB)? Exogenous fatty acids induce a pro-inflammatory cytokine response in human placental cells in vitro via activation of TLR4 signaling pathways. The placenta is exposed to changes in circulating maternal fatty acid concentrations throughout pregnancy. Fatty acids are master regulators of innate immune pathways through recruitment of toll-like receptors and activation of cytokine synthesis. Trophoblast cells isolated from 14 normal term human placentas were incubated with long chain fatty acids (FA) of different carbon length and degree of saturation. The expression and secretion of interleukin-6 (IL-6), IL-8 and tumor necrosis factor-alpha (TNF-α) were measured by reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Antibodies against TLR4 ligand binding domain, downstream signaling and anti-p65 NFkB-inhibitor were used to characterize the pathways of FA action. General approach used primary human term trophoblast cell culture. Methods and end-points used real-time quantitative PCR, cytokine measurements, immunohistochemistry, western blots. The long chain saturated fatty acids, stearic and palmitic (PA), stimulated the synthesis as well as the release of TNF-α, IL-6 and IL-8 by trophoblast cells (2- to 6-fold, P acids did not modify cytokine expression significantly. Palmitate-induced inflammatory effects were mediated via TLR4 activation, NF-kB phosphorylation and nuclear translocation. TNF-α protein level was close to the limit of detection in the culture medium even when cells were cultured with PA. These mechanisms open the way to a better understanding of how changes in maternal lipid homeostasis may regulate placental inflammatory status. X.Y. was recipient of fellowship award from West China Second University Hospital, Sichuan University (NIH HD 22965-19). The authors have nothing

Kynurenine acid - metabolism and regulation of kynurenine pathway

Directory of Open Access Journals (Sweden)

Piotr Kozłowski

2017-07-01

Full Text Available Kynurenic acid (KYNA was first isolated from the dog's urine in 1853 by german chemist Justus von Liebig. KYNA probably plays an important role in the pathogenesis of many neurodegenerative and psychiatric diseases. Its elevated concentration were found in the brain (post mortem or in the cerebrospinal fluid patients  with schizophrenia, bipolar disorder, Alzheimer's disease, meningitis, autoimmune diseases, inflammatory processes and memory and learning disorders. The reduced KYNA concentration is characteristic for multiple sclerosis, Parkinson's disease, Huntington's disease and epilepsy. KYNA is an organic compound naturally occurring in nature. This amino acid belongs to the group of exogenous amino acids and can be synthesized by plants and bacteria alone. The largest amount of tryptophan about 95%is  metabolised by the kynurenine pathway. Only 1% of tryptophan supplied in the diet serves to produce serotonin in the brain. The process of regulation of KYNA synthesis in both the CNS and the periphery is complicated.

The role of leucine in isoprenoid metabolism. Incorporation of [3-13C]leucine and of [2-3H,4-14C]-β,β-dimethyl-acrylic acid into phytosterols by tissue cultures of Andrographis paniculata

International Nuclear Information System (INIS)

Anastasis, P.; Freer, I.; Overton, K.; Rycroft, D.; Singh, S.B.

1985-01-01

[3- 13 C]Leucine is incorporated into phytosterols by tissue cultures of Andrographis paniculata by breakdown to acetyl-CoA and its subsequent incorporation via (3S)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) and mevalonic acid; [2- 3 H,4- 14 C]-β,β-dimethylacrylic acid also is not incorporated intact. (author)

Biosynthesis of sesquiterpenes in grape berry exocarp of Vitis vinifera L.: evidence for a transport of farnesyl diphosphate precursors from plastids to the cytosol.

Science.gov (United States)

May, Bianca; Lange, B Markus; Wüst, Matthias

2013-11-01

The participation of the mevalonic acid (MVA) and 1-deoxy-d-xylulose 5-phosphate/2-C-methyl-d-erythritol-4-phosphate (DOXP/MEP) pathways in sesquiterpene biosynthesis of grape berries was investigated. There is an increasing interest in this class of terpenoids, since the oxygenated sesquiterpene rotundone was identified as the peppery aroma impact compound in Australian Shiraz wines. To investigate precursor supply pathway utilization, in vivo feeding experiments were performed with the deuterium labeled, pathway specific, precursors [5,5-(2)H2]-1-deoxy-d-xylulose and [5,5-(2)H2]-mevalonic acid lactone. Head Space-Solid Phase Micro Extraction-Gas Chromatography-Mass Spectrometry (HS-SPME-GC-MS) analysis of the generated volatile metabolites demonstrated that de novo sesquiterpene biosynthesis is mainly located in the grape berry exocarp (skin), with no detectable activity in the mesocarp (flesh) of the Lemberger variety. Interestingly, precursors from both the (primarily) cytosolic MVA and plastidial DOXP/MEP pathways were incorporated into grape sesquiterpenes in the varieties Lemberger, Gewürztraminer and Syrah. Our labeling data provide evidence for a homogenous, cytosolic pool of precursors for sesquiterpene biosynthesis, indicating that a transport of precursors occurs mostly from plastids to the cytosol. The labeling patterns of the sesquiterpene germacrene D were in agreement with a cyclization mechanism analogous to that of a previously cloned enantioselective (R)-germacrene D synthase from Solidago canadensis. This observation was subsequently confirmed by enantioselective GC-MS analysis demonstrating the exclusive presence of (R)-germacrene D, and not the (S)-enantiomer, in grape berries. Copyright © 2013 Elsevier Ltd. All rights reserved.

Transcriptome analysis of bitter acid biosynthesis and precursor pathways in hop (Humulus lupulus

Directory of Open Access Journals (Sweden)

Clark Shawn M

2013-01-01

Full Text Available Abstract Background Bitter acids (e.g. humulone are prenylated polyketides synthesized in lupulin glands of the hop plant (Humulus lupulus which are important contributors to the bitter flavour and stability of beer. Bitter acids are formed from acyl-CoA precursors derived from branched-chain amino acid (BCAA degradation and C5 prenyl diphosphates from the methyl-D-erythritol 4-phosphate (MEP pathway. We used RNA sequencing (RNA-seq to obtain the transcriptomes of isolated lupulin glands, cones with glands removed and leaves from high α-acid hop cultivars, and analyzed these datasets for genes involved in bitter acid biosynthesis including the supply of major precursors. We also measured the levels of BCAAs, acyl-CoA intermediates, and bitter acids in glands, cones and leaves. Results Transcripts encoding all the enzymes of BCAA metabolism were significantly more abundant in lupulin glands, indicating that BCAA biosynthesis and subsequent degradation occurs in these specialized cells. Branched-chain acyl-CoAs and bitter acids were present at higher levels in glands compared with leaves and cones. RNA-seq analysis showed the gland-specific expression of the MEP pathway, enzymes of sucrose degradation and several transcription factors that may regulate bitter acid biosynthesis in glands. Two branched-chain aminotransferase (BCAT enzymes, HlBCAT1 and HlBCAT2, were abundant, with gene expression quantification by RNA-seq and qRT-PCR indicating that HlBCAT1 was specific to glands while HlBCAT2 was present in glands, cones and leaves. Recombinant HlBCAT1 and HlBCAT2 catalyzed forward (biosynthetic and reverse (catabolic reactions with similar kinetic parameters. HlBCAT1 is targeted to mitochondria where it likely plays a role in BCAA catabolism. HlBCAT2 is a plastidial enzyme likely involved in BCAA biosynthesis. Phylogenetic analysis of the hop BCATs and those from other plants showed that they group into distinct biosynthetic (plastidial and

Transcriptome mining and in silico structural and functional analysis of ascorbic acid and tartaric acid biosynthesis pathway enzymes in rose-scanted geranium.

Science.gov (United States)

Narnoliya, Lokesh K; Sangwan, Rajender S; Singh, Sudhir P

2018-06-01

Rose-scented geranium (Pelargonium sp.) is widely known as aromatic and medicinal herb, accumulating specialized metabolites of high economic importance, such as essential oils, ascorbic acid, and tartaric acid. Ascorbic acid and tartaric acid are multifunctional metabolites of human value to be used as vital antioxidants and flavor enhancing agents in food products. No information is available related to the structural and functional properties of the enzymes involved in ascorbic acid and tartaric acid biosynthesis in rose-scented geranium. In the present study, transcriptome mining was done to identify full-length genes, followed by their bioinformatic and molecular modeling investigations and understanding of in silico structural and functional properties of these enzymes. Evolutionary conserved domains were identified in the pathway enzymes. In silico physicochemical characterization of the catalytic enzymes revealed isoelectric point (pI), instability index, aliphatic index, and grand average hydropathy (GRAVY) values of the enzymes. Secondary structural prediction revealed abundant proportion of alpha helix and random coil confirmations in the pathway enzymes. Three-dimensional homology models were developed for these enzymes. The predicted structures showed significant structural similarity with their respective templates in root mean square deviation analysis. Ramachandran plot analysis of the modeled enzymes revealed that more than 84% of the amino acid residues were within the favored regions. Further, functionally important residues were identified corresponding to catalytic sites located in the enzymes. To, our best knowledge, this is the first report which provides a foundation on functional annotation and structural determination of ascorbic acid and tartaric acid pathway enzymes in rose-scanted geranium.

Wnt/β-catenin signalling pathway mediated aberrant hippocampal neurogenesis in kainic acid-induced epilepsy.

Science.gov (United States)

Qu, Zhengyi; Su, Fang; Qi, Xueting; Sun, Jianbo; Wang, Hongcai; Qiao, Zhenkui; Zhao, Hong; Zhu, Yulan

2017-10-01

Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis with massive neuronal loss and severe gliosis. Aberrant neurogenesis has been shown in the epileptogenesis process of temporal lobe epilepsy. However, the molecular mechanisms underlying aberrant neurogenesis remain unclear. The roles of Wnt signalling cascade have been well established in neurogenesis during multiple aspects. Here, we used kainic acid-induced rat epilepsy model to investigate whether Wnt/β-catenin signalling pathway is involved in the aberrant neurogenesis in temporal lobe epilepsy. Immunostaining and western blotting results showed that the expression levels of β-catenin, Wnt3a, and cyclin D1, the key regulators in Wnt signalling pathway, were up-regulated during acute epilepsy induced by the injection of kainic acids, indicating that Wnt signalling pathway was activated in kainic acid-induced temporal lobe epilepsy. Moreover, BrdU labelling results showed that blockade of the Wnt signalling by knocking down β-catenin attenuated aberrant neurogenesis induced by kainic acids injection. Altogether, Wnt/β-catenin signalling pathway mediated hippocampal neurogenesis during epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Temporal lobe epilepsy is a chronic disorder of nerve system, mainly characterized by hippocampal sclerosis. Aberrant neurogenesis has been shown to involve in the epileptogenesis process of temporal lobe epilepsy. In the present study, we discovered that Wnt3a/β-catenin signalling pathway serves as a link between aberrant neurogenesis and underlying remodelling in the hippocampus, leading to temporal lobe epilepsy, which might provide new strategies for clinical treatment of temporal lobe epilepsy. Copyright © 2017 John Wiley & Sons, Ltd.

Rewiring the reductive tricarboxylic acid pathway and L-malate transport pathway of Aspergillus oryzae for overproduction of L-malate.

Science.gov (United States)

Liu, Jingjing; Xie, Zhipeng; Shin, Hyun-Dong; Li, Jianghua; Du, Guocheng; Chen, Jian; Liu, Long

2017-07-10

Aspergillus oryzae finds wide application in the food, feed, and wine industries, and is an excellent cell factory platform for production of organic acids. In this work, we achieved the overproduction of L-malate by rewiring the reductive tricarboxylic acid (rTCA) pathway and L-malate transport pathway of A. oryzae NRRL 3488. First, overexpression of native pyruvate carboxylase and malate dehydrogenase in the rTCA pathway improved the L-malate titer from 26.1gL -1 to 42.3gL -1 in shake flask culture. Then, the oxaloacetate anaplerotic reaction was constructed by heterologous expression of phosphoenolpyruvate carboxykinase and phosphoenolpyruvate carboxylase from Escherichia coli, increasing the L-malate titer to 58.5gL -1 . Next, the export of L-malate from the cytoplasm to the external medium was strengthened by overexpression of a C4-dicarboxylate transporter gene from A. oryzae and an L-malate permease gene from Schizosaccharomyces pombe, improving the L-malate titer from 58.5gL -1 to 89.5gL -1 . Lastly, guided by transcription analysis of the expression profile of key genes related to L-malate synthesis, the 6-phosphofructokinase encoded by the pfk gene was identified as a potential limiting step for L-malate synthesis. Overexpression of pfk with the strong sodM promoter increased the L-malate titer to 93.2gL -1 . The final engineered A. oryzae strain produced 165gL -1 L-malate with a productivity of 1.38gL -1 h -1 in 3-L fed-batch culture. Overall, we constructed an efficient L-malate producer by rewiring the rTCA pathway and L-malate transport pathway of A. oryzae NRRL 3488, and the engineering strategy adopted here may be useful for the construction of A. oryzae cell factories to produce other organic acids. Copyright © 2017 Elsevier B.V. All rights reserved.

Role of leucine in isoprenoid metabolism. Incorporation of (3-/sup 13/C)leucine and of (2-/sup 3/H,4-/sup 14/C)-. beta. ,. beta. -dimethyl-acrylic acid into phytosterols by tissue cultures of Andrographis paniculata

Energy Technology Data Exchange (ETDEWEB)

Anastasis, P; Freer, I; Overton, K; Rycroft, D; Singh, S B [Glasgow Univ. (UK). Dept. of Chemistry

1985-02-01

(3-/sup 13/C)Leucine is incorporated into phytosterols by tissue cultures of Andrographis paniculata by breakdown to acetyl-CoA and its subsequent incorporation via (3S)-3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) and mevalonic acid; (2-/sup 3/H,4-/sup 14/C)-..beta..,..beta..-dimethylacrylic acid also is not incorporated intact.

Agmatine deiminase pathway genes in Lactobacillus brevis are linked to the tyrosine decarboxylation operon in a putative acid resistance locus

NARCIS (Netherlands)

Lucas, Patrick M.; Blancato, Victor S.; Claisse, Olivier; Magni, Christian; Lolkema, Juke S.; Lonvaud-Funel, Aline

In lactic acid bacteria (LAB), amino acids and their derivatives may be converted into amine-containing compounds designated biogenic amines, in pathways providing metabolic energy and/ or acid resistance to the bacteria. In a previous study, a pathway converting tyrosine to tyramine was detected in

Functions and Signaling Pathways of Amino Acids in Intestinal Inflammation

Directory of Open Access Journals (Sweden)

Fang He

2018-01-01

Full Text Available Intestine is always exposed to external environment and intestinal microorganism; thus it is more sensitive to dysfunction and dysbiosis, leading to intestinal inflammation, such as inflammatory bowel disease (IBD, irritable bowel syndrome (IBS, and diarrhea. An increasing number of studies indicate that dietary amino acids play significant roles in preventing and treating intestinal inflammation. The review aims to summarize the functions and signaling mechanisms of amino acids in intestinal inflammation. Amino acids, including essential amino acids (EAAs, conditionally essential amino acids (CEAAs, and nonessential amino acids (NEAAs, improve the functions of intestinal barrier and expressions of anti-inflammatory cytokines and tight junction proteins but decrease oxidative stress and the apoptosis of enterocytes as well as the expressions of proinflammatory cytokines in the intestinal inflammation. The functions of amino acids are associated with various signaling pathways, including mechanistic target of rapamycin (mTOR, inducible nitric oxide synthase (iNOS, calcium-sensing receptor (CaSR, nuclear factor-kappa-B (NF-κB, mitogen-activated protein kinase (MAPK, nuclear erythroid-related factor 2 (Nrf2, general controlled nonrepressed kinase 2 (GCN2, and angiotensin-converting enzyme 2 (ACE2.

Necrotrophic pathogens use the salicylic acid signaling pathway to promote disease development in tomato.

Science.gov (United States)

Rahman, Taha Abd El; Oirdi, Mohamed El; Gonzalez-Lamothe, Rocio; Bouarab, Kamal

2012-12-01

Plants use different immune pathways to combat pathogens. The activation of the jasmonic acid (JA)-signaling pathway is required for resistance against necrotrophic pathogens; however, to combat biotrophic pathogens, the plants activate mainly the salicylic acid (SA)-signaling pathway. SA can antagonize JA signaling and vice versa. NPR1 (noninducible pathogenesis-related 1) is considered a master regulator of SA signaling. NPR1 interacts with TGA transcription factors, ultimately leading to the activation of SA-dependent responses. SA has been shown to promote disease development caused by the necrotrophic pathogen Botrytis cinerea through NPR1, by suppressing the expression of two JA-dependent defense genes, proteinase inhibitors I and II. We show here that the transcription factor TGA1.a contributes to disease development caused by B. cinerea in tomato by suppressing the expression of proteinase inhibitors I and II. Finally, we present evidence that the SA-signaling pathway contributes to disease development caused by another necrotrophic pathogen, Alternaria solani, in tomato. Disease development promoted by SA through NPR1 requires the TGA1.a transcription factor. These data highlight how necrotrophs manipulate the SAsignaling pathway to promote their disease in tomato.

Sterol Biosynthesis Pathway as Target for Anti-trypanosomatid Drugs

Directory of Open Access Journals (Sweden)

Wanderley de Souza

2009-01-01

Full Text Available Sterols are constituents of the cellular membranes that are essential for their normal structure and function. In mammalian cells, cholesterol is the main sterol found in the various membranes. However, other sterols predominate in eukaryotic microorganisms such as fungi and protozoa. It is now well established that an important metabolic pathway in fungi and in members of the Trypanosomatidae family is one that produces a special class of sterols, including ergosterol, and other 24-methyl sterols, which are required for parasitic growth and viability, but are absent from mammalian host cells. Currently, there are several drugs that interfere with sterol biosynthesis (SB that are in use to treat diseases such as high cholesterol in humans and fungal infections. In this review, we analyze the effects of drugs such as (a statins, which act on the mevalonate pathway by inhibiting HMG-CoA reductase, (b bisphosphonates, which interfere with the isoprenoid pathway in the step catalyzed by farnesyl diphosphate synthase, (c zaragozic acids and quinuclidines, inhibitors of squalene synthase (SQS, which catalyzes the first committed step in sterol biosynthesis, (d allylamines, inhibitors of squalene epoxidase, (e azoles, which inhibit C14α-demethylase, and (f azasterols, which inhibit Δ24(25-sterol methyltransferase (SMT. Inhibition of this last step appears to have high selectivity for fungi and trypanosomatids, since this enzyme is not found in mammalian cells. We review here the IC50 values of these various inhibitors, their effects on the growth of trypanosomatids (both in axenic cultures and in cell cultures, and their effects on protozoan structural organization (as evaluted by light and electron microscopy and lipid composition. The results show that the mitochondrial membrane as well as the membrane lining the protozoan cell body and flagellum are the main targets. Probably as a consequence of these primary effects, other important changes take

Targeting Cytosolic Nucleic Acid-Sensing Pathways for Cancer Immunotherapies.

Science.gov (United States)

Iurescia, Sandra; Fioretti, Daniela; Rinaldi, Monica

2018-01-01

The innate immune system provides the first line of defense against pathogen infection though also influences pathways involved in cancer immunosurveillance. The innate immune system relies on a limited set of germ line-encoded sensors termed pattern recognition receptors (PRRs), signaling proteins and immune response factors. Cytosolic receptors mediate recognition of danger damage-associated molecular patterns (DAMPs) signals. Once activated, these sensors trigger multiple signaling cascades, converging on the production of type I interferons and proinflammatory cytokines. Recent studies revealed that PRRs respond to nucleic acids (NA) released by dying, damaged, cancer cells, as danger DAMPs signals, and presence of signaling proteins across cancer types suggests that these signaling mechanisms may be involved in cancer biology. DAMPs play important roles in shaping adaptive immune responses through the activation of innate immune cells and immunological response to danger DAMPs signals is crucial for the host response to cancer and tumor rejection. Furthermore, PRRs mediate the response to NA in several vaccination strategies, including DNA immunization. As route of double-strand DNA intracellular entry, DNA immunization leads to expression of key components of cytosolic NA-sensing pathways. The involvement of NA-sensing mechanisms in the antitumor response makes these pathways attractive drug targets. Natural and synthetic agonists of NA-sensing pathways can trigger cell death in malignant cells, recruit immune cells, such as DCs, CD8 + T cells, and NK cells, into the tumor microenvironment and are being explored as promising adjuvants in cancer immunotherapies. In this minireview, we discuss how cGAS-STING and RIG-I-MAVS pathways have been targeted for cancer treatment in preclinical translational researches. In addition, we present a targeted selection of recent clinical trials employing agonists of cytosolic NA-sensing pathways showing how these pathways

Up-regulation of abscisic acid signaling pathway facilitates aphid xylem absorption and osmoregulation under drought stress.

Science.gov (United States)

Guo, Huijuan; Sun, Yucheng; Peng, Xinhong; Wang, Qinyang; Harris, Marvin; Ge, Feng

2016-02-01

The activation of the abscisic acid (ABA) signaling pathway reduces water loss from plants challenged by drought stress. The effect of drought-induced ABA signaling on the defense and nutrition allocation of plants is largely unknown. We postulated that these changes can affect herbivorous insects. We studied the effects of drought on different feeding stages of pea aphids in the wild-type A17 of Medicago truncatula and ABA signaling pathway mutant sta-1. We examined the impact of drought on plant water status, induced plant defense signaling via the abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA) pathways, and on the host nutritional quality in terms of leaf free amino acid content. During the penetration phase of aphid feeding, drought decreased epidermis/mesophyll resistance but increased mesophyll/phloem resistance of A17 but not sta-1 plants. Quantification of transcripts associated with ABA, JA and SA signaling indicated that the drought-induced up-regulation of ABA signaling decreased the SA-dependent defense but increased the JA-dependent defense in A17 plants. During the phloem-feeding phase, drought had little effect on the amino acid concentrations and the associated aphid phloem-feeding parameters in both plant genotypes. In the xylem absorption stage, drought decreased xylem absorption time of aphids in both genotypes because of decreased water potential. Nevertheless, the activation of the ABA signaling pathway increased water-use efficiency of A17 plants by decreasing the stomatal aperture and transpiration rate. In contrast, the water potential of sta-1 plants (unable to close stomata) was too low to support xylem absorption activity of aphids; the aphids on sta-1 plants had the highest hemolymph osmolarity and lowest abundance under drought conditions. Taken together this study illustrates the significance of cross-talk between biotic-abiotic signaling pathways in plant-aphid interaction, and reveals the mechanisms leading to alter

Dual biosynthetic pathways to phytosterol via cycloartenol and lanosterol in Arabidopsis.

Science.gov (United States)

Ohyama, Kiyoshi; Suzuki, Masashi; Kikuchi, Jun; Saito, Kazuki; Muranaka, Toshiya

2009-01-20

The differences between the biosynthesis of sterols in higher plants and yeast/mammals are believed to originate at the cyclization step of oxidosqualene, which is cyclized to cycloartenol in higher plants and lanosterol in yeast/mammals. Recently, lanosterol synthase genes were identified from dicotyledonous plant species including Arabidopsis, suggesting that higher plants possess dual biosynthetic pathways to phytosterols via lanosterol, and through cycloartenol. To identify the biosynthetic pathway to phytosterol via lanosterol, and to reveal the contributions to phytosterol biosynthesis via each cycloartenol and lanosterol, we performed feeding experiments by using [6-(13)C(2)H(3)]mevalonate with Arabidopsis seedlings. Applying (13)C-{(1)H}{(2)H} nuclear magnetic resonance (NMR) techniques, the elucidation of deuterium on C-19 behavior of phytosterol provided evidence that small amounts of phytosterol were biosynthesized via lanosterol. The levels of phytosterol increased on overexpression of LAS1, and phytosterols derived from lanosterol were not observed in a LAS1-knockout plant. This is direct evidence to indicate that the biosynthetic pathway for phytosterol via lanosterol exists in plant cells. We designate the biosynthetic pathway to phytosterols via lanosterol "the lanosterol pathway." LAS1 expression is reported to be induced by the application of jasmonate and is thought to have evolved from an ancestral cycloartenol synthase to a triterpenoid synthase, such as beta-amyrin synthase and lupeol synthase. Considering this background, the lanosterol pathway may contribute to the biosynthesis of not only phytosterols, but also steroids as secondary metabolites.

Stimulation of the Salicylic Acid Pathway Aboveground Recruits Entomopathogenic Nematodes Belowground.

Directory of Open Access Journals (Sweden)

Camila Cramer Filgueiras

Full Text Available Plant defense pathways play a critical role in mediating tritrophic interactions between plants, herbivores, and natural enemies. While the impact of plant defense pathway stimulation on natural enemies has been extensively explored aboveground, belowground ramifications of plant defense pathway stimulation are equally important in regulating subterranean pests and still require more attention. Here we investigate the effect of aboveground stimulation of the salicylic acid pathway through foliar application of the elicitor methyl salicylate on belowground recruitment of the entomopathogenic nematode, Steinernema diaprepesi. Also, we implicate a specific root-derived volatile that attracts S. diaprepesi belowground following aboveground plant stimulation by an elicitor. In four-choice olfactometer assays, citrus plants treated with foliar applications of methyl salicylate recruited S. diaprepesi in the absence of weevil feeding as compared with negative controls. Additionally, analysis of root volatile profiles of citrus plants receiving foliar application of methyl salicylate revealed production of d-limonene, which was absent in negative controls. The entomopathogenic nematode S. diaprepesi was recruited to d-limonene in two-choice olfactometer trials. These results reinforce the critical role of plant defense pathways in mediating tritrophic interactions, suggest a broad role for plant defense pathway signaling belowground, and hint at sophisticated plant responses to pest complexes.

Stimulation of the Salicylic Acid Pathway Aboveground Recruits Entomopathogenic Nematodes Belowground

Science.gov (United States)

Filgueiras, Camila Cramer; Willett, Denis S.; Junior, Alcides Moino; Pareja, Martin; Borai, Fahiem El; Dickson, Donald W.; Stelinski, Lukasz L.; Duncan, Larry W.

2016-01-01

Plant defense pathways play a critical role in mediating tritrophic interactions between plants, herbivores, and natural enemies. While the impact of plant defense pathway stimulation on natural enemies has been extensively explored aboveground, belowground ramifications of plant defense pathway stimulation are equally important in regulating subterranean pests and still require more attention. Here we investigate the effect of aboveground stimulation of the salicylic acid pathway through foliar application of the elicitor methyl salicylate on belowground recruitment of the entomopathogenic nematode, Steinernema diaprepesi. Also, we implicate a specific root-derived volatile that attracts S. diaprepesi belowground following aboveground plant stimulation by an elicitor. In four-choice olfactometer assays, citrus plants treated with foliar applications of methyl salicylate recruited S. diaprepesi in the absence of weevil feeding as compared with negative controls. Additionally, analysis of root volatile profiles of citrus plants receiving foliar application of methyl salicylate revealed production of d-limonene, which was absent in negative controls. The entomopathogenic nematode S. diaprepesi was recruited to d-limonene in two-choice olfactometer trials. These results reinforce the critical role of plant defense pathways in mediating tritrophic interactions, suggest a broad role for plant defense pathway signaling belowground, and hint at sophisticated plant responses to pest complexes. PMID:27136916

Phenylalanine ammonia lyase catalyzed synthesis of amino acids by an MIO-cofactor independent pathway.

Science.gov (United States)

Lovelock, Sarah L; Lloyd, Richard C; Turner, Nicholas J

2014-04-25

Phenylalanine ammonia lyases (PALs) belong to a family of 4-methylideneimidazole-5-one (MIO) cofactor dependent enzymes which are responsible for the conversion of L-phenylalanine into trans-cinnamic acid in eukaryotic and prokaryotic organisms. Under conditions of high ammonia concentration, this deamination reaction is reversible and hence there is considerable interest in the development of PALs as biocatalysts for the enantioselective synthesis of non-natural amino acids. Herein the discovery of a previously unobserved competing MIO-independent reaction pathway, which proceeds in a non-stereoselective manner and results in the generation of both L- and D-phenylalanine derivatives, is described. The mechanism of the MIO-independent pathway is explored through isotopic-labeling studies and mutagenesis of key active-site residues. The results obtained are consistent with amino acid deamination occurring by a stepwise E1 cB elimination mechanism. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The effect of O-acetylsalicylic acid on lipid synthesis by guinea pig gastric mucosa in vitro

International Nuclear Information System (INIS)

Spohn, M.; McColl, I.

1987-01-01

The aim of this work was to investigate the involvement of lipids as possible components of the gastric mucosal barrier by studying the synthesis and secretion of lipids by the epithelial cell lining of gastric mucosa and the effect of salicylate on these processes. O-Acetylsalicylic acid reversibly reduced in vitro incorporation of (U- 14 C) and of DL-(2- 14 C) mevalonic acid into lipids by isolated epithelial cells and by intact mucosa of guinea pig stomach, indicating reversible inhibition of lipid synthesis by the tissue in the presence of the drug. Inhibition of incorporation of both precursors into total lipids, into their fatty acid components, and into cholesterol is demonstrated

Distribution of δ-aminolevulinic acid biosynthetic pathways among phototrophic and related bacteria

International Nuclear Information System (INIS)

Avissar, Y.J.; Beale, S.I.; Ormerod, J.G.

1989-01-01

Two biosynthetic pathways are known for the universal tetrapyrrole precursor, δ-aminolevulinic acid (ALA): condensation of glycine and succinyl-CoA to form ALA with the loss of C-1 of glycine as CO 2 , and conversion of the intact carbon skeleton of glutamate to ALA in a process requiring tRNA Glu , ATP, Mg 2+ , NADPH, and pyridoxal phosphate. The distribution of the two ALA biosynthetic pathways among various bacterial genera was determined, using cell-free extracts obtained from representative organisms. Evidence for the operation of the glutamate pathway was obtained by the measurement of RNase-sensitive label incorporation from glutamate into ALA using 3,4-[ 3 H]glutamate and 1-[ 14 C]glutamate as substrate. The glycine pathway was indicated by RNase-insensitive incorporation of level from 2-[ 14 C]glycine into ALA. The distribution of the two pathways among the bacteria tested was in general agreement with their previously phylogenetic relationships and clearly indicates that the glutamate pathway is the more ancient process, whereas the glycine pathway probably evolved much later. The glutamate pathway is the more widely utilized one among bacteria, while the glycine pathway is apparently limited to the α subgroup of purple bacteria (including Rhodobacter, Rhodospirillum, and Rhizobium). E. coli was found ALA via the glutamate pathway. The ALA-requiring hemA mutant of E. coli was determined to lack the dehydrogenase activity that utilizes glutamyl-tRNA as a substrate

FPPS mediates TGF-β1-induced non-small cell lung cancer cell invasion and the EMT process via the RhoA/Rock1 pathway.

Science.gov (United States)

Lin, Lin; Li, Ming; Lin, Lei; Xu, Xiaolin; Jiang, Gening; Wu, Liang

2018-02-05

Farnesyl pyrophosphate synthase (FPPS), a key enzyme in the mevalonate pathway, was recently shown to play a role in cancer progression. However, its role in non-small cell lung cancer (NSCLC) metastasis and the underlying mechanism remain unclear. In this study, FPPS expression was significantly correlated with TNM stage, and metastasis. Inhibition or knockdown of FPPS blocked TGF-β1-induced cell invasion and epithelial-to-mesenchymal transition (EMT) process. FPPS expression of FPPS was induced by TGF-β1 and FPPS promoted cell invasion and EMT via the RhoA/Rock1 pathway. In conclusion, FPPS mediates TGF-β1-induced lung cancer cell invasion and EMT via the RhoA/Rock1 pathway. These findings suggest new treatment strategies to reduce mortality associated with metastasis in patients with NSCLC. Copyright © 2018 Elsevier Inc. All rights reserved.

Pachymic acid promotes induction of autophagy related to IGF-1 signaling pathway in WI-38 cells.

Science.gov (United States)

Lee, Su-Gyeong; Kim, Moon-Moo

2017-12-01

The insulin-like growth factor 1 (IGF-1) signaling pathway has spotlighted as a mechanism to elucidate aging associated with autophagy in recent years. Therefore, we have tried to screen an effective compound capable of inducing autophagy to delay aging process. The aim of this study is to investigate whether pachymic acid, a main compound in Poria cocos, induces autophagy in the aged cells. The aging of young cells was induced by treatment with IGF-1 at 50 ng/ml three times every two days. The effect of pachymic acid on cell viability was evaluated in human lung fibroblasts, WI-38 cells, using MTT assay. The induction of autophagy was detected using autophagy detection kit. The expression of proteins related to autophagy and IGF-1 signaling pathway was examined by western blot analysis and immunofluorescence assay. In this study, pachymic acid showed cytotoxic effect in a dose dependent manner and remarkably induced autophagy at the same time. Moreover, pachymic acid increased the expression of proteins related to autophagy such as LC3-II and Beclin1 and decreased the levels of mTor phosphorylation and p70S6K in the aged cells. In particular, pachymic acid increased the expression of p-PI3K, p-FoxO and Catalase. In addition, pachymic acid remarkably increased the expression of IGFBP-3. Above results suggest that pachymic acid could induce autophagy related to IGF-1 signaling pathway in the aged cells. Copyright © 2017 Elsevier GmbH. All rights reserved.

Porting the synthetic D-glucaric acid pathway from Escherichia coli to Saccharomyces cerevisiae.

Science.gov (United States)

Gupta, Amita; Hicks, Michael A; Manchester, Shawn P; Prather, Kristala L J

2016-09-01

D-Glucaric acid can be produced as a value-added chemical from biomass through a de novo pathway in Escherichia coli. However, previous studies have identified pH-mediated toxicity at product concentrations of 5 g/L and have also found the eukaryotic myo-inositol oxygenase (MIOX) enzyme to be rate-limiting. We ported this pathway to Saccaromyces cerevisiae, which is naturally acid-tolerant and evaluate a codon-optimized MIOX homologue. We constructed two engineered yeast strains that were distinguished solely by their MIOX gene - either the previous version from Mus musculus or a homologue from Arabidopsis thaliana codon-optimized for expression in S. cerevisiae - in order to identify the rate-limiting steps for D-glucaric acid production both from a fermentative and non-fermentative carbon source. myo-Inositol availability was found to be rate-limiting from glucose in both strains and demonstrated to be dependent on growth rate, whereas the previously used M. musculus MIOX activity was found to be rate-limiting from glycerol. Maximum titers were 0.56 g/L from glucose in batch mode, 0.98 g/L from glucose in fed-batch mode, and 1.6 g/L from glucose supplemented with myo-inositol. Future work focusing on the MIOX enzyme, the interplay between growth and production modes, and promoting aerobic respiration should further improve this pathway. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Monoclonal Antibodies Follow Distinct Aggregation Pathways During Production-Relevant Acidic Incubation and Neutralization

DEFF Research Database (Denmark)

Pedersen, Thomas Skamris; Tian, Xinsheng; Thorolfsson, Matthias

2016-01-01

and orthogonal analytical methods, including small-angle X-ray scattering and dynamic light scattering and supplemented the experimental data with crystal structure-based spatial aggregation propensity (SAP) calculations. RESULTS: We revealed distinct solution behaviors between the three mAb models: At acidic p......PURPOSE: Aggregation aspects of therapeutic monoclonal antibodies (mAbs) are of common concern to the pharmaceutical industry. Low pH treatment is applied during affinity purification and to inactivate endogenous retroviruses, directing interest to the mechanisms of acid-induced antibody...... distinguish between reversible and irreversible mAb aggregation pathways at early stages of acidic treatment....

Compound-Specific Isotopic Analysis of Meteoritic Amino Acids as a Tool for Evaluating Potential Formation Pathways

Science.gov (United States)

Elsila, Jamie E.; Burton, Aaron S.; Callahan, Michael C.; Charnley, Steven B.; Glavin, Daniel P.; Dworkin, Jason P.

2012-01-01

Measurements of stable hydrogen, carbon, and nitrogen isotopic ratios (delta D, delta C-13, delta N-15) of organic compounds can reveal information about their origin and formation pathways. Several formation mechanisms and environments have been postulated for the amino acids detected in carbonaceous chondrites. As each proposed mechanism utilizes different precursor molecules, the isotopic signatures of the resulting amino acids may point towards the most likely of these proposed pathways. The technique of gas chromatography coupled with mass spectrometry and isotope ratio mass spectrometry provides compound-specific structural and isotopic information from a single splitless injection, enhancing the amount of information gained from small amounts of precious samples such as carbonaceous chondrites. We have applied this technique to measure the compound-specific C, N, and H isotopic ratios of amino acids from seven CM and CR carbonaceous chondrites. We are using these measurements to evaluate predictions of expected isotopic enrichments from potential formation pathways and environments, leading to a better understanding of the origin of these compounds.

Mechanisms of action and medicinal applications of abscisic Acid.

Science.gov (United States)

Bassaganya-Riera, J; Skoneczka, J; Kingston, D G J; Krishnan, A; Misyak, S A; Guri, A J; Pereira, A; Carter, A B; Minorsky, P; Tumarkin, R; Hontecillas, R

2010-01-01

Since its discovery in the early 1960's, abscisic acid (ABA) has received considerable attention as an important phytohormone, and more recently, as a candidate medicinal in humans. In plants it has been shown to regulate important physiological processes such as response to drought stress, and dormancy. The discovery of ABA synthesis in animal cells has generated interest in the possible parallels between its role in plant and animal systems. The importance of this molecule has prompted the development of several methods for the chemical synthesis of ABA, which differ significantly from the biosynthesis of ABA in plants through the mevalonic acid pathway. ABA recognition in plants has been shown to occur at both the intra- and extracellularly but little is known about the perception of ABA by animal cells. A few ABA molecular targets have been identified in vitro (e.g., calcium signaling, G protein-coupled receptors) in both plant and animal systems. A unique finding in mammalian systems, however, is that the peroxisome proliferator-activated receptor, PPAR gamma, is upregulated by ABA in both in vitro and in vivo studies. Comparison of the human PPAR gamma gene network with Arabidopsis ABA-related genes reveal important orthologs between these groups. Also, ABA can ameliorate the symptoms of type II diabetes, targeting PPAR gamma in a similar manner as the thiazolidinediones class of anti-diabetic drugs. The use of ABA in the treatment of type II diabetes, offers encouragement for further studies concerning the biomedical applications of ABA.

Molecular cloning and functional expression of geranylgeranyl pyrophosphate synthase from Coleus forskohlii Briq

Directory of Open Access Journals (Sweden)

Kawamukai Makoto

2004-11-01

Full Text Available Abstract Background Isopentenyl diphosphate (IPP, a common biosynthetic precursor to the labdane diterpene forskolin, has been biosynthesised via a non-mevalonate pathway. Geranylgeranyl diphosphate (GGPP synthase is an important branch point enzyme in terpenoid biosynthesis. Therefore, GGPP synthase is thought to be a key enzyme in biosynthesis of forskolin. Herein we report the first confirmation of the GGPP synthase gene in Coleus forskohlii Briq. Results The open reading frame for full-length GGPP synthase encodes a protein of 359 amino acids, in which 1,077 nucleotides long with calculated molecular mass of 39.3 kDa. Alignments of C. forskohlii GGPP synthase amino acid sequences revealed high homologies with other plant GGPP synthases. Several highly conserved regions, including two aspartate-rich motifs were identified. Transient expression of the N-terminal region of C. forskohlii GGPP synthase-GFP fusion protein in tobacco cells demonstrated subcellular localization in the chloroplast. Carotenoid production was observed in Escherichia coli harboring pACCAR25ΔcrtE from Erwinia uredovora and plasmid carrying C. forskohlii GGPP synthase. These results suggested that cDNA encoded functional GGPP synthase. Furthermore, C. forskohlii GGPP synthase expression was strong in leaves, decreased in stems and very little expression was observed in roots. Conclusion This investigation proposed that forskolin was synthesised via a non-mevalonate pathway. GGPP synthase is thought to be involved in the biosynthesis of forskolin, which is primarily synthesised in the leaves and subsequently accumulates in the stems and roots.

Metabolic pathways regulated by abscisic acid, salicylic acid and γ-aminobutyric acid in association with improved drought tolerance in creeping bentgrass (Agrostis stolonifera).

Science.gov (United States)

Li, Zhou; Yu, Jingjin; Peng, Yan; Huang, Bingru

2017-01-01

Abscisic acid (ABA), salicylic acid (SA) and γ-aminobutyric acid (GABA) are known to play roles in regulating plant stress responses. This study was conducted to determine metabolites and associated pathways regulated by ABA, SA and GABA that could contribute to drought tolerance in creeping bentgrass (Agrostis stolonifera). Plants were foliar sprayed with ABA (5 μM), GABA (0.5 mM) and SA (10 μM) or water (untreated control) prior to 25 days drought stress in controlled growth chambers. Application of ABA, GABA or SA had similar positive effects on alleviating drought damages, as manifested by the maintenance of lower electrolyte leakage and greater relative water content in leaves of treated plants relative to the untreated control. Metabolic profiling showed that ABA, GABA and SA induced differential metabolic changes under drought stress. ABA mainly promoted the accumulation of organic acids associated with tricarboxylic acid cycle (aconitic acid, succinic acid, lactic acid and malic acid). SA strongly stimulated the accumulation of amino acids (proline, serine, threonine and alanine) and carbohydrates (glucose, mannose, fructose and cellobiose). GABA enhanced the accumulation of amino acids (GABA, glycine, valine, proline, 5-oxoproline, serine, threonine, aspartic acid and glutamic acid) and organic acids (malic acid, lactic acid, gluconic acid, malonic acid and ribonic acid). The enhanced drought tolerance could be mainly due to the enhanced respiration metabolism by ABA, amino acids and carbohydrates involved in osmotic adjustment (OA) and energy metabolism by SA, and amino acid metabolism related to OA and stress-defense secondary metabolism by GABA. © 2016 Scandinavian Plant Physiology Society.

Amino acids and insulin act additively to regulate components of the ubiquitin-proteasome pathway in C2C12 myotubes

Directory of Open Access Journals (Sweden)

Lomax Michael A

2007-03-01

Full Text Available Abstract Background The ubiquitin-proteasome system is the predominant pathway for myofibrillar proteolysis but a previous study in C2C12 myotubes only observed alterations in lysosome-dependent proteolysis in response to complete starvation of amino acids or leucine from the media. Here, we determined the interaction between insulin and amino acids in the regulation of myotube proteolysis Results Incubation of C2C12 myotubes with 0.2 × physiological amino acids concentration (0.2 × PC AA, relative to 1.0 × PC AA, significantly increased total proteolysis and the expression of 14-kDa E2 ubiquitin conjugating enzyme (p Conclusion In a C2C12 myotube model of myofibrillar protein turnover, amino acid limitation increases proteolysis in a ubiquitin-proteasome-dependent manner. Increasing amino acids or leucine alone, act additively with insulin to down regulate proteolysis and expression of components of ubiquitin-proteasome pathway. The effects of amino acids on proteolysis but not insulin and leucine, are blocked by inhibition of the mTOR signalling pathway.

Role of plastidic pyruvate dehydrogenase complex (pl. PDC) in chloroplast metabolism of spinach

International Nuclear Information System (INIS)

Schulze-Siebert, D.; Homeyer, U.; Schultz, G.

1986-01-01

Labeling experiments of chloroplasts in the light ( 14 CO 2 , 2- 14 C-pyruvate etc.) revealed that pl. PDC is predominantly involved in the synthesis of branched chain amino acids and pl. isoprenoids (carotenes, PQ, α-T). In this context, pl. phosphoglycerate mutase as missing link in the C 3 → C 2 metabolism of chloroplasts was identified by latency experiments. This indicates a direct pathway from Calvin cycle to pl. PDC. Using protoplasts, maximal rates in pl. PDC metabolism were obtained. On the other hand, mitochondrial PDC in protoplasts is mainly involved in fatty acid synthesis by known mechanism. Additionally, cytosolic-ER-isoprenoids were formed (e.g. sterols). When 14 CO 2 was simultaneously applied with unlabeled acetate to protoplasts in the light an isotopic dilution of fatty acids were found but not of pl. isoprenoids. This may indicate an partially channeling of pl. PDC and mevalonate pathway for pl. isoprenoid synthesis. Inhibitory studies with DCMU point in the same direction

Fatty Acid Biosynthesis Pathways in Methylomicrobium buryatense 5G(B1).

Science.gov (United States)

Demidenko, Aleksandr; Akberdin, Ilya R; Allemann, Marco; Allen, Eric E; Kalyuzhnaya, Marina G

2016-01-01

Methane utilization by methanotrophic bacteria is an attractive application for biotechnological conversion of natural or biogas into high-added-value products. Haloalcaliphilic methanotrophic bacteria belonging to the genus Methylomicrobium are among the most promising strains for methane-based biotechnology, providing easy and inexpensive cultivation, rapid growth, and the availability of established genetic tools. A number of methane bioconversions using these microbial cultures have been discussed, including the derivation of biodiesel, alkanes, and OMEGA-3 supplements. These compounds are derived from bacterial fatty acid pools. Here, we investigate fatty acid biosynthesis in Methylomicrobium buryatense 5G(B1) . Most of the genes homologous to typical Type II fatty acid biosynthesis pathways could be annotated by bioinformatics analyses, with the exception of fatty acid transport and regulatory elements. Different approaches for improving fatty acid accumulation were investigated. These studies indicated that both fatty acid degradation and acetyl- and malonyl-CoA levels are bottlenecks for higher level fatty acid production. The best strain generated in this study synthesizes 111 ± 2 mg/gDCW of extractable fatty acids, which is ~20% more than the original strain. A candidate gene for fatty acid biosynthesis regulation, farE , was identified and studied. Its deletion resulted in drastic changes to the fatty acid profile, leading to an increased pool of C18-fatty acid methyl ester. The FarE-regulon was further investigated by RNA-seq analysis of gene expression in farE -knockout mutants and farE -overexpressing strains. These gene profiles highlighted a novel set of enzymes and regulators involved in fatty acid biosynthesis. The gene expression and fatty acid profiles of the different farE -strains support the hypothesis that metabolic fluxes upstream of fatty acid biosynthesis restrict fatty acid production in the methanotroph.

Changes in actin dynamics are involved in salicylic acid signaling pathway.

Science.gov (United States)

Matoušková, Jindřiška; Janda, Martin; Fišer, Radovan; Sašek, Vladimír; Kocourková, Daniela; Burketová, Lenka; Dušková, Jiřina; Martinec, Jan; Valentová, Olga

2014-06-01

Changes in actin cytoskeleton dynamics are one of the crucial players in many physiological as well as non-physiological processes in plant cells. Positioning of actin filament arrays is necessary for successful establishment of primary lines of defense toward pathogen attack, depolymerization leads very often to the enhanced susceptibility to the invading pathogen. On the other hand it was also shown that the disruption of actin cytoskeleton leads to the induction of defense response leading to the expression of PATHOGENESIS RELATED proteins (PR). In this study we show that pharmacological actin depolymerization leads to the specific induction of genes in salicylic acid pathway but not that involved in jasmonic acid signaling. Life imaging of leafs of Arabidopsis thaliana with GFP-tagged fimbrin (GFP-fABD2) treated with 1 mM salicylic acid revealed rapid disruption of actin filaments resembling the pattern viewed after treatment with 200 nM latrunculin B. The effect of salicylic acid on actin filament fragmentation was prevented by exogenous addition of phosphatidic acid, which binds to the capping protein and thus promotes actin polymerization. The quantitative evaluation of actin filament dynamics is also presented. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

The shikimate pathway: review of amino acid sequence, function and three-dimensional structures of the enzymes.

Science.gov (United States)

Mir, Rafia; Jallu, Shais; Singh, T P

2015-06-01

The aromatic compounds such as aromatic amino acids, vitamin K and ubiquinone are important prerequisites for the metabolism of an organism. All organisms can synthesize these aromatic metabolites through shikimate pathway, except for mammals which are dependent on their diet for these compounds. The pathway converts phosphoenolpyruvate and erythrose 4-phosphate to chorismate through seven enzymatically catalyzed steps and chorismate serves as a precursor for the synthesis of variety of aromatic compounds. These enzymes have shown to play a vital role for the viability of microorganisms and thus are suggested to present attractive molecular targets for the design of novel antimicrobial drugs. This review focuses on the seven enzymes of the shikimate pathway, highlighting their primary sequences, functions and three-dimensional structures. The understanding of their active site amino acid maps, functions and three-dimensional structures will provide a framework on which the rational design of antimicrobial drugs would be based. Comparing the full length amino acid sequences and the X-ray crystal structures of these enzymes from bacteria, fungi and plant sources would contribute in designing a specific drug and/or in developing broad-spectrum compounds with efficacy against a variety of pathogens.

Salicylic acid induces vanillin synthesis through the phospholipid signaling pathway in Capsicum chinense cell cultures.

Science.gov (United States)

Rodas-Junco, Beatriz A; Cab-Guillén, Yahaira; Muñoz-Sánchez, J Armando; Vázquez-Flota, Felipe; Monforte-González, Miriam; Hernández-Sotomayor, S M Teresa

2013-10-01

Signal transduction via phospholipids is mediated by phospholipases such as phospholipase C (PLC) and D (PLD), which catalyze hydrolysis of plasma membrane structural phospholipids. Phospholipid signaling is also involved in plant responses to phytohormones such as salicylic acid (SA). The relationships between phospholipid signaling, SA, and secondary metabolism are not fully understood. Using a Capsicum chinense cell suspension as a model, we evaluated whether phospholipid signaling modulates SA-induced vanillin production through the activation of phenylalanine ammonia lyase (PAL), a key enzyme in the biosynthetic pathway. Salicylic acid was found to elicit PAL activity and consequently vanillin production, which was diminished or reversed upon exposure to the phosphoinositide-phospholipase C (PI-PLC) signaling inhibitors neomycin and U73122. Exposure to the phosphatidic acid inhibitor 1-butanol altered PLD activity and prevented SA-induced vanillin production. Our results suggest that PLC and PLD-generated secondary messengers may be modulating SA-induced vanillin production through the activation of key biosynthetic pathway enzymes.

Perturbations in amino acids and metabolic pathways in osteoarthritis patients determined by targeted metabolomics analysis.

Science.gov (United States)

Chen, Rui; Han, Su; Liu, Xuefeng; Wang, Kunpeng; Zhou, Yong; Yang, Chundong; Zhang, Xi

2018-05-15

Osteoarthritis (OA) is a degenerative synovial joint disease affecting people worldwide. However, the exact pathogenesis of OA remains unclear. Metabolomics analysis was performed to obtain insight into possible pathogenic mechanisms and diagnostic biomarkers of OA. Ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-TQ-MS), followed by multivariate statistical analysis, was used to determine the serum amino acid profiles of 32 OA patients and 35 healthy controls. Variable importance for project values and Student's t-test were used to determine the metabolic abnormalities in OA. Another 30 OA patients were used as independent samples to validate the alterations in amino acids. MetaboAnalyst was used to identify the key amino acid pathways and construct metabolic networks describing their relationships. A total of 25 amino acids and four biogenic amines were detected by UPLC-TQ-MS. Differences in amino acid profiles were found between the healthy controls and OA patients. Alanine, γ-aminobutyric acid and 4-hydroxy-l-proline were important biomarkers distinguishing OA patients from healthy controls. The metabolic pathways with the most significant effects were involved in metabolism of alanine, aspartate, glutamate, arginine and proline. The results of this study improve understanding of the amino acid metabolic abnormalities and pathogenic mechanisms of OA at the molecular level. The metabolic perturbations may be important for the diagnosis and prevention of OA. Copyright © 2018 Elsevier B.V. All rights reserved.

Enhancement of arachidonic acid signaling pathway by nicotinic acid receptor HM74A

International Nuclear Information System (INIS)

Tang, Yuting; Zhou, Lubing; Gunnet, Joseph W.; Wines, Pamela G.; Cryan, Ellen V.; Demarest, Keith T.

2006-01-01

HM74A is a G protein-coupled receptor for nicotinic acid (niacin), which has been used clinically to treat dyslipidemia for decades. The molecular mechanisms whereby niacin exerts its pleiotropic effects on lipid metabolism remain largely unknown. In addition, the most common side effect in niacin therapy is skin flushing that is caused by prostaglandin release, suggesting that the phospholipase A 2 (PLA 2 )/arachidonic acid (AA) pathway is involved. Various eicosanoids have been shown to activate peroxisome-proliferator activated receptors (PPAR) that play a diverse array of roles in lipid metabolism. To further elucidate the potential roles of HM74A in mediating the therapeutic effects and/or side effects of niacin, we sought to explore the signaling events upon HM74A activation. Here we demonstrated that HM74A synergistically enhanced UTP- and bradykinin-mediated AA release in a pertussis toxin-sensitive manner in A431 cells. Activation of HM74A also led to Ca 2+ -mobilization and enhanced bradykinin-promoted Ca 2+ -mobilization through Gi protein. While HM74A increased ERK1/2 activation by the bradykinin receptor, it had no effects on UTP-promoted ERK1/2 activation.Furthermore, UTP- and bradykinin-mediated AA release was significantly decreased in the presence of both MAPK kinase inhibitor PD 098059 and PKC inhibitor GF 109203X. However, the synergistic effects of HM74A were not dramatically affected by co-treatment with both inhibitors, indicating the cross-talk occurred at the receptor level. Finally, stimulation of A431 cells transiently transfected with PPRE-luciferase with AA significantly induced luciferase activity, mimicking the effects of PPARγ agonist rosiglitazone, suggesting that alteration of AA signaling pathway can regulate gene expression via endogenous PPARs

Enhancement of arachidonic acid signaling pathway by nicotinic acid receptor HM74A

Energy Technology Data Exchange (ETDEWEB)

Tang, Yuting [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States); Zhou, Lubing [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States); Gunnet, Joseph W [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States); Wines, Pamela G [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States); Cryan, Ellen V [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States); Demarest, Keith T [Endocrine Therapeutics and Metabolic Disorders, Johnson and Johnson Pharmaceutical Research and Development, L.L.C., 1000 Rt. 202, Raritan, NJ 08869 (United States)

2006-06-23

HM74A is a G protein-coupled receptor for nicotinic acid (niacin), which has been used clinically to treat dyslipidemia for decades. The molecular mechanisms whereby niacin exerts its pleiotropic effects on lipid metabolism remain largely unknown. In addition, the most common side effect in niacin therapy is skin flushing that is caused by prostaglandin release, suggesting that the phospholipase A{sub 2} (PLA{sub 2})/arachidonic acid (AA) pathway is involved. Various eicosanoids have been shown to activate peroxisome-proliferator activated receptors (PPAR) that play a diverse array of roles in lipid metabolism. To further elucidate the potential roles of HM74A in mediating the therapeutic effects and/or side effects of niacin, we sought to explore the signaling events upon HM74A activation. Here we demonstrated that HM74A synergistically enhanced UTP- and bradykinin-mediated AA release in a pertussis toxin-sensitive manner in A431 cells. Activation of HM74A also led to Ca{sup 2+}-mobilization and enhanced bradykinin-promoted Ca{sup 2+}-mobilization through Gi protein. While HM74A increased ERK1/2 activation by the bradykinin receptor, it had no effects on UTP-promoted ERK1/2 activation.Furthermore, UTP- and bradykinin-mediated AA release was significantly decreased in the presence of both MAPK kinase inhibitor PD 098059 and PKC inhibitor GF 109203X. However, the synergistic effects of HM74A were not dramatically affected by co-treatment with both inhibitors, indicating the cross-talk occurred at the receptor level. Finally, stimulation of A431 cells transiently transfected with PPRE-luciferase with AA significantly induced luciferase activity, mimicking the effects of PPAR{gamma} agonist rosiglitazone, suggesting that alteration of AA signaling pathway can regulate gene expression via endogenous PPARs.

A Novel Antibiotic Mechanism of l-Cyclopropylalanine Blocking the Biosynthetic Pathway of Essential Amino Acid l-Leucine

Directory of Open Access Journals (Sweden)

Bingji Ma

2017-12-01

Full Text Available The unusual amino acid l-cyclopropylalanine was isolated from the mushroom Amanita virgineoides after detection in an anti-fungal screening test. l-Cyclopropylalanine was found to exhibit broad-spectrum inhibition against fungi and bacteria. The anti-fungal activity was found to be abolished in the presence of the amino acid l-leucine, but not any other amino acids, indicating that l-cyclopropylalanine may block the biosynthesis of the essential amino acid l-leucine, thereby inhibiting fungal and bacteria growth. Further biochemical studies found l-cyclopropylalanine indeed inhibits α-isopropylmalate synthase (α-IMPS, the enzyme that catalyzes the rate-limiting step in the biosynthetic pathway of l-leucine. Inhibition of essential l-leucine synthesis in fungal and bacteria organisms, a pathway absent in host organisms such as humans, may represent a novel antibiotic mechanism to counter the ever-increasing problem of drug resistance to existing antibiotics.

Ellagic acid radiosensitizes tumor cells by evoking apoptotic pathway

International Nuclear Information System (INIS)

Ahire, Vidhula R.; Mishra, K.P.

2016-01-01

Cancer causes millions of deaths each year globally. In most patients, the cause of treatment failure is found associated with the resistance to chemotherapy and radiotherapy. The development of tumor cell resistance evokes multiple intracellular molecular pathways. In addition, the limitation in treatment outcome arises due to unintended cytotoxic effects of the synthetic anticancer drugs to normal cells and tissues. Considerable focus of research is, therefore, devoted to examine plant-based herbal compounds which may prove potential anticancer drug for developing effective cancer therapy. Research results from our laboratory have shown that ellagic acid (EA), a natural flavonoid displays enhanced tumor toxicity in combination with gamma radiation to many types of cancers in vitro as well as in vivo. Studies on the underlying mechanisms of toxicity suggest that EA employs the cellular signaling pathways in producing the observed effects. This paper gives an account of molecular mechanisms of EA-induced apoptosis process in tumor cytotoxicity. It is suggested that EA acts as a novel radiosensitizer for tumors and a radioprotector for normal cells which may offer a novel protocol for cancer treatment. (author)

N2O production pathways in the subtropical acid forest soils in China

International Nuclear Information System (INIS)

Zhang Jinbo; Cai Zucong; Zhu Tongbin

2011-01-01

To date, N 2 O production pathways are poorly understood in the humid subtropical and tropical forest soils. A 15 N-tracing experiment was carried out under controlled laboratory conditions to investigate the processes responsible for N 2 O production in four subtropical acid forest soils (pH 2 O emission in the subtropical acid forest soils, being responsible for 56.1%, 53.5%, 54.4%, and 55.2% of N 2 O production, in the GC, GS, GB, and TC soils, respectively, under aerobic conditions (40%-52%WFPS). The heterotrophic nitrification (recalcitrant organic N oxidation) accounted for 27.3%-41.8% of N 2 O production, while the contribution of autotrophic nitrification was little in the studied subtropical acid forest soils. The ratios of N 2 O-N emission from total nitrification (heterotrophic+autotrophic nitrification) were higher than those in most previous references. The soil with the lowest pH and highest organic-C content (GB) had the highest ratio (1.63%), suggesting that soil pH-organic matter interactions may exist and affect N 2 O product ratios from nitrification. The ratio of N 2 O-N emission from heterotrophic nitrification varied from 0.02% to 25.4% due to soil pH and organic matter. Results are valuable in the accurate modeling of N2O production in the subtropical acid forest soils and global budget. - Highlights: → We studied N 2 O production pathways in subtropical acid forest soil under aerobic conditions. → Denitrification was the main source of N 2 O production in subtropical acid forest soils. → Heterotrophic nitrification accounted for 27.3%-41.8% of N 2 O production. → While, contribution of autotrophic nitrification to N 2 O production was little. → Ratios of N 2 O-N emission from nitrification were higher than those in most previous references.

Oral benfotiamine plus alpha-lipoic acid normalises complication-causing pathways in type 1 diabetes.

Science.gov (United States)

Du, X; Edelstein, D; Brownlee, M

2008-10-01

We determined whether fixed doses of benfotiamine in combination with slow-release alpha-lipoic acid normalise markers of reactive oxygen species-induced pathways of complications in humans. Male participants with and without type 1 diabetes were studied in the General Clinical Research Centre of the Albert Einstein College of Medicine. Glycaemic status was assessed by measuring baseline values of three different indicators of hyperglycaemia. Intracellular AGE formation, hexosamine pathway activity and prostacyclin synthase activity were measured initially, and after 2 and 4 weeks of treatment. In the nine participants with type 1 diabetes, treatment had no effect on any of the three indicators used to assess hyperglycaemia. However, treatment with benfotiamine plus alpha-lipoic acid completely normalised increased AGE formation, reduced increased monocyte hexosamine-modified proteins by 40% and normalised the 70% decrease in prostacyclin synthase activity from 1,709 +/- 586 pg/ml 6-keto-prostaglandin F(1alpha) to 4,696 +/- 533 pg/ml. These results show that the previously demonstrated beneficial effects of these agents on complication-causing pathways in rodent models of diabetic complications also occur in humans with type 1 diabetes.

De Novo Biosynthesis of Glutarate via α-Keto Acid Carbon Chain Extension and Decarboxylation Pathway in Escherichia coli.

Science.gov (United States)

Wang, Jian; Wu, Yifei; Sun, Xinxiao; Yuan, Qipeng; Yan, Yajun

2017-10-20

Microbial based bioplastics are promising alternatives to petroleum based synthetic plastics due to their renewability and economic feasibility. Glutarate is one of the most potential building blocks for bioplastics. The recent biosynthetic routes for glutarate were mostly based on the l-lysine degradation pathway from Pseudomonas putida that required lysine either by feeding or lysine overproduction via genetic manipulations. Herein, we established a novel glutarate biosynthetic pathway by incorporation of a "+1" carbon chain extension pathway from α-ketoglutarate (α-KG) in combination with α-keto acid decarboxylation pathway in Escherichia coli. Introduction of homocitrate synthase (HCS), homoaconitase (HA) and homoisocitrate dehydrogenase (HICDH) from Saccharomyces cerevisiae into E. coli enabled "+1" carbon extension from α-KG to α-ketoadipate (α-KA), which was subsequently converted into glutarate by a promiscuous α-keto acid decarboxylase (KivD) and a succinate semialdehyde dehydrogenase (GabD). The recombinant E. coli coexpressing all five genes produced 0.3 g/L glutarate from glucose. To further improve the titers, α-KG was rechanneled into carbon chain extension pathway via the clustered regularly interspersed palindromic repeats system mediated interference (CRISPRi) of essential genes sucA and sucB in tricarboxylic acid (TCA) cycle. The final strain could produce 0.42 g/L glutarate, which was increased by 40% compared with the parental strain.

Mitochondrial localization of the mevalonate pathway enzyme 3-Hydroxy-3-methyl-glutaryl-CoA reductase in the Trypanosomatidae

DEFF Research Database (Denmark)

Pena Diaz, Javier; Montalvetti, Andrea; Flores, Carmen-Lisset

2004-01-01

3-Hydroxy-3-methyl-glutaryl-CoA reductase (HMGR) is a key enzyme in the sterol biosynthesis pathway, but its subcellular distribution in the Trypanosomatidae family is somewhat controversial. Trypanosoma cruzi and Leishmania HMGRs are closely related in their catalytic domains to bacterial and eu...

Uric acid priming in human monocytes is driven by the AKT–PRAS40 autophagy pathway

Science.gov (United States)

Crişan, Tania O.; Cleophas, Maartje C. P.; Novakovic, Boris; Erler, Kathrin; van de Veerdonk, Frank L.; Stunnenberg, Hendrik G.; Netea, Mihai G.; Dinarello, Charles A.; Joosten, Leo A. B.

2017-01-01

Metabolic triggers are important inducers of the inflammatory processes in gout. Whereas the high serum urate levels observed in patients with gout predispose them to the formation of monosodium urate (MSU) crystals, soluble urate also primes for inflammatory signals in cells responding to gout-related stimuli, but also in other common metabolic diseases. In this study, we investigated the mechanisms through which uric acid selectively lowers human blood monocyte production of the natural inhibitor IL-1 receptor antagonist (IL-1Ra) and shifts production toward the highly inflammatory IL-1β. Monocytes from healthy volunteers were first primed with uric acid for 24 h and then subjected to stimulation with lipopolysaccharide (LPS) in the presence or absence of MSU. Transcriptomic analysis revealed broad inflammatory pathways associated with uric acid priming, with NF-κB and mammalian target of rapamycin (mTOR) signaling strongly increased. Functional validation did not identify NF-κB or AMP-activated protein kinase phosphorylation, but uric acid priming induced phosphorylation of AKT and proline-rich AKT substrate 40 kDa (PRAS 40), which in turn activated mTOR. Subsequently, Western blot for the autophagic structure LC3-I and LC3-II (microtubule-associated protein 1A/1B-light chain 3) fractions, as well as fluorescence microscopy of LC3-GFP–overexpressing HeLa cells, revealed lower autophagic activity in cells exposed to uric acid compared with control conditions. Interestingly, reactive oxygen species production was diminished by uric acid priming. Thus, the Akt–PRAS40 pathway is activated by uric acid, which inhibits autophagy and recapitulates the uric acid-induced proinflammatory cytokine phenotype. PMID:28484006

Uric acid priming in human monocytes is driven by the AKT-PRAS40 autophagy pathway.

Science.gov (United States)

Crişan, Tania O; Cleophas, Maartje C P; Novakovic, Boris; Erler, Kathrin; van de Veerdonk, Frank L; Stunnenberg, Hendrik G; Netea, Mihai G; Dinarello, Charles A; Joosten, Leo A B

2017-05-23

Metabolic triggers are important inducers of the inflammatory processes in gout. Whereas the high serum urate levels observed in patients with gout predispose them to the formation of monosodium urate (MSU) crystals, soluble urate also primes for inflammatory signals in cells responding to gout-related stimuli, but also in other common metabolic diseases. In this study, we investigated the mechanisms through which uric acid selectively lowers human blood monocyte production of the natural inhibitor IL-1 receptor antagonist (IL-1Ra) and shifts production toward the highly inflammatory IL-1β. Monocytes from healthy volunteers were first primed with uric acid for 24 h and then subjected to stimulation with lipopolysaccharide (LPS) in the presence or absence of MSU. Transcriptomic analysis revealed broad inflammatory pathways associated with uric acid priming, with NF-κB and mammalian target of rapamycin (mTOR) signaling strongly increased. Functional validation did not identify NF-κB or AMP-activated protein kinase phosphorylation, but uric acid priming induced phosphorylation of AKT and proline-rich AKT substrate 40 kDa (PRAS 40), which in turn activated mTOR. Subsequently, Western blot for the autophagic structure LC3-I and LC3-II (microtubule-associated protein 1A/1B-light chain 3) fractions, as well as fluorescence microscopy of LC3-GFP-overexpressing HeLa cells, revealed lower autophagic activity in cells exposed to uric acid compared with control conditions. Interestingly, reactive oxygen species production was diminished by uric acid priming. Thus, the Akt-PRAS40 pathway is activated by uric acid, which inhibits autophagy and recapitulates the uric acid-induced proinflammatory cytokine phenotype.

Impairment of PPARα and the Fatty Acid Oxidation Pathway Aggravates Renal Fibrosis during Aging.

Science.gov (United States)

Chung, Ki Wung; Lee, Eun Kyeong; Lee, Mi Kyung; Oh, Goo Taeg; Yu, Byung Pal; Chung, Hae Young

2018-04-01

Defects in the renal fatty acid oxidation (FAO) pathway have been implicated in the development of renal fibrosis. Although, compared with young kidneys, aged kidneys show significantly increased fibrosis with impaired kidney function, the mechanisms underlying the effects of aging on renal fibrosis have not been investigated. In this study, we investigated peroxisome proliferator-activated receptor α (PPAR α ) and the FAO pathway as regulators of age-associated renal fibrosis. The expression of PPAR α and the FAO pathway-associated proteins significantly decreased with the accumulation of lipids in the renal tubular epithelial region during aging in rats. In particular, decreased PPAR α protein expression associated with increased expression of PPAR α -targeting microRNAs. Among the microRNAs with increased expression during aging, miR-21 efficiently decreased PPAR α expression and impaired FAO when ectopically expressed in renal epithelial cells. In cells pretreated with oleic acid to induce lipid stress, miR-21 treatment further enhanced lipid accumulation. Furthermore, treatment with miR-21 significantly exacerbated the TGF- β -induced fibroblast phenotype of epithelial cells. We verified the physiologic importance of our findings in a calorie restriction model. Calorie restriction rescued the impaired FAO pathway during aging and slowed fibrosis development. Finally, compared with kidneys of aged littermate controls, kidneys of aged PPAR α -/- mice showed exaggerated lipid accumulation, with decreased activity of the FAO pathway and a severe fibrosis phenotype. Our results suggest that impaired renal PPAR α signaling during aging aggravates renal fibrosis development, and targeting PPAR α is useful for preventing age-associated CKD. Copyright © 2018 by the American Society of Nephrology.

Enhancement of carotenoid production by disrupting the C22-sterol desaturase gene (CYP61 in Xanthophyllomyces dendrorhous

Directory of Open Access Journals (Sweden)

Loto Iris

2012-10-01

Full Text Available Abstract Background Xanthophyllomyces dendrorhous is a basidiomycetous yeast that synthesizes astaxanthin, which is a carotenoid with a great biotechnological impact. The ergosterol and carotenoid synthesis pathways are derived from the mevalonate pathway, and in both pathways, cytochrome P450 enzymes are involved. Results In this study, we isolated and described the X. dendrorhous CYP61 gene, which encodes a cytochrome P450 involved in ergosterol biosynthesis. This gene is composed of nine exons and encodes a 526 amino acid polypeptide that shares significant percentages of identity and similitude with the C22-sterol desaturase, CYP61, from other fungi. Mutants derived from different parental strains were obtained by disrupting the CYP61 gene with an antibiotic selection marker. These mutants were not able to produce ergosterol and accumulated ergosta-5,8,22-trien-3-ol and ergosta-5,8-dien-3-ol. Interestingly, all of the mutants had a more intense red color phenotype than their respective parental strains. The carotenoid composition was qualitatively and quantitatively analyzed by RP-HPLC, revealing that the carotenoid content was higher in the mutant strains without major changes in their composition. The expression of the HMGR gene, which encodes an enzyme involved in the mevalonate pathway (3-hydroxy-3-methylglutaryl-CoA reductase, was analyzed by RT-qPCR showing that its transcript levels are higher in the CYP61 mutants. Conclusions These results suggest that in X. dendrorhous, ergosterol regulates HMGR gene expression by a negative feedback mechanism and in this way; it contributes in the regulation of the carotenoid biosynthesis.

Study of Arachidonic Acid Pathway in Human Bladder Tumor

Directory of Open Access Journals (Sweden)

Masahide Matsuyama

2009-12-01

Full Text Available Recent epidemiological studies and animal experiments have demonstrated that nonsteroidal anti-inflammatory drugs (NSAIDs reduce the incidence of colorectal carcinoma. Cyclooxygenase (COX is the principal target of NSAIDs. COX is the first oxidase in the process of prostaglandin production from arachidonic acid. COX enzyme may be involved in the initiation and/or the promotion of tumorigenesis due to NSAIDs inhibition of COX. Lipoxygenase (LOX is also an initial enzyme in the pathway for producing leukotrienes from arachidonic acid. Similar to COX, LOX enzyme may also be involved in the initiation and/or promotion of tumorigenesis. Peroxisome proliferator activator-receptor (PPAR-γ is a ligand-activated transcriptional factor belonging to the steroid receptor superfamily. PPAR-γ plays a role in both adipocyte differentiation and tumorigenesis. PPAR-γ is one target for cell growth modulation of NSAIDs. In this review, we report the expression of COX-2, LOX and PPAR-γ in human bladder tumor tissues as well as the effects of COX-2 and LOX inhibitors and PPAR-γ ligand.

Study of Arachidonic Acid Pathway in Human Bladder Tumor

Directory of Open Access Journals (Sweden)

Masahide Matsuyama

2009-01-01

Full Text Available Recent epidemiological studies and animal experiments have demonstrated that nonsteroidal anti-inflammatory drugs (NSAIDs reduce the incidence of colorectal carcinoma. Cyclooxygenase (COX is the principal target of NSAIDs. COX is the first oxidase in the process of prostaglandin production from arachidonic acid. COX enzyme may be involved in the initiation and/or the promotion of tumorigenesis due to NSAIDs inhibition of COX. Lipoxygenase (LOX is also an initial enzyme in the pathway for producing leukotrienes from arachidonic acid. Similar to COX, LOX enzyme may also be involved in the initiation and/or promotion of tumorigenesis. Peroxisome proliferator activator-receptor (PPAR-γ is a ligand-activated transcriptional factor belonging to the steroid receptor superfamily. PPAR-γ plays a role in both adipocyte differentiation and tumorigenesis. PPAR-γ is one target for cell growth modulation of NSAIDs. In this review, we report the expression of COX-2, LOX and PPAR-γ in human bladder tumor tissues as well as the effects of COX-2 and LOX inhibitors and PPAR-γ ligand.

A Newly Discovered Antifibrotic Pathway Regulated by Two Fatty Acid Receptors: GPR40 and GPR84.

Science.gov (United States)

Gagnon, Lyne; Leduc, Martin; Thibodeau, Jean-Francois; Zhang, Ming-Zhi; Grouix, Brigitte; Sarra-Bournet, Francois; Gagnon, William; Hince, Kathy; Tremblay, Mikaël; Geerts, Lilianne; Kennedy, Christopher R J; Hébert, Richard L; Gutsol, Alex; Holterman, Chet E; Kamto, Eldjonai; Gervais, Liette; Ouboudinar, Jugurtha; Richard, Jonathan; Felton, Alexandra; Laverdure, Alexandre; Simard, Jean-Christophe; Létourneau, Sylvie; Cloutier, Marie-Pier; Leblond, Francois A; Abbott, Shaun D; Penney, Christopher; Duceppe, Jean-Simon; Zacharie, Boulos; Dupuis, Jocelyn; Calderone, Angelino; Nguyen, Quang T; Harris, Raymond C; Laurin, Pierre

2018-05-01

Numerous clinical conditions can lead to organ fibrosis and functional failure. There is a great need for therapies that could effectively target pathophysiological pathways involved in fibrosis. GPR40 and GPR84 are G protein-coupled receptors with free fatty acid ligands and are associated with metabolic and inflammatory disorders. Although GPR40 and GPR84 are involved in diverse physiological processes, no evidence has demonstrated the relevance of GPR40 and GPR84 in fibrosis pathways. Using PBI-4050 (3-pentylbenzeneacetic acid sodium salt), a synthetic analog of a medium-chain fatty acid that displays agonist and antagonist ligand affinity toward GPR40 and GPR84, respectively, we uncovered an antifibrotic pathway involving these receptors. In experiments using Gpr40- and Gpr84-knockout mice in models of kidney fibrosis (unilateral ureteral obstruction, long-term post-acute ischemic injury, and adenine-induced chronic kidney disease), we found that GPR40 is protective and GPR84 is deleterious in these diseases. Moreover, through binding to GPR40 and GPR84, PBI-4050 significantly attenuated fibrosis in many injury contexts, as evidenced by the antifibrotic activity observed in kidney, liver, heart, lung, pancreas, and skin fibrosis models. Therefore, GPR40 and GPR84 may represent promising molecular targets in fibrosis pathways. We conclude that PBI-4050 is a first-in-class compound that may be effective for managing inflammatory and fibrosis-related diseases. Copyright © 2018 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Fatty acid hydroperoxides pathways in plants. A review.

Directory of Open Access Journals (Sweden)

Fauconnier, M. L.

1997-02-01

Full Text Available The present paper focusses on the fatty acid hydroperoxides pathways, mainly hydroperoxide lyase and hydroperoxide dehydrase. For each enzyme, the definition, occurrence and subcellular localization is presented. Particular attention is given to reaction mecanisms and to substrate specificity. Physiological roles of reaction products are also discussed.

El presente artículo se centra en las rutas de los hidroperóxidos de ácidos grasos, principalmente la hidroperóxido liasa y la hidroperóxido dehidrasa. Se presenta para cada enzima, la definición, distribución y localización subcelular. Se da atención particular a los mecanismos de reacción y a la especificidad de sustrato. También se discuten los papeles fisiológicos de los productos de reacción.

Analysis of hydroxycinnamic acid degradation in Agrobacterium fabrum reveals a coenzyme A-dependent, beta-oxidative deacetylation pathway.

Science.gov (United States)

Campillo, Tony; Renoud, Sébastien; Kerzaon, Isabelle; Vial, Ludovic; Baude, Jessica; Gaillard, Vincent; Bellvert, Floriant; Chamignon, Cécile; Comte, Gilles; Nesme, Xavier; Lavire, Céline; Hommais, Florence

2014-06-01

The soil- and rhizosphere-inhabiting bacterium Agrobacterium fabrum (genomospecies G8 of the Agrobacterium tumefaciens species complex) is known to have species-specific genes involved in ferulic acid degradation. Here, we characterized, by genetic and analytical means, intermediates of degradation as feruloyl coenzyme A (feruloyl-CoA), 4-hydroxy-3-methoxyphenyl-β-hydroxypropionyl-CoA, 4-hydroxy-3-methoxyphenyl-β-ketopropionyl-CoA, vanillic acid, and protocatechuic acid. The genes atu1416, atu1417, and atu1420 have been experimentally shown to be necessary for the degradation of ferulic acid. Moreover, the genes atu1415 and atu1421 have been experimentally demonstrated to be essential for this degradation and are proposed to encode a phenylhydroxypropionyl-CoA dehydrogenase and a 4-hydroxy-3-methoxyphenyl-β-ketopropionic acid (HMPKP)-CoA β-keto-thiolase, respectively. We thus demonstrated that the A. fabrum hydroxycinnamic degradation pathway is an original coenzyme A-dependent β-oxidative deacetylation that could also transform p-coumaric and caffeic acids. Finally, we showed that this pathway enables the metabolism of toxic compounds from plants and their use for growth, likely providing the species an ecological advantage in hydroxycinnamic-rich environments, such as plant roots or decaying plant materials.

Reconstruction of cytosolic fumaric acid biosynthetic pathways in Saccharomyces cerevisiae

Directory of Open Access Journals (Sweden)

Xu Guoqiang

2012-02-01

Full Text Available Abstract Background Fumaric acid is a commercially important component of foodstuffs, pharmaceuticals and industrial materials, yet the current methods of production are unsustainable and ecologically destructive. Results In this study, the fumarate biosynthetic pathway involving reductive reactions of the tricarboxylic acid cycle was exogenously introduced in S. cerevisiae by a series of simple genetic modifications. First, the Rhizopus oryzae genes for malate dehydrogenase (RoMDH and fumarase (RoFUM1 were heterologously expressed. Then, expression of the endogenous pyruvate carboxylase (PYC2 was up-regulated. The resultant yeast strain, FMME-001 ↑PYC2 + ↑RoMDH, was capable of producing significantly higher yields of fumarate in the glucose medium (3.18 ± 0.15 g liter-1 than the control strain FMME-001 empty vector. Conclusions The results presented here provide a novel strategy for fumarate biosynthesis, which represents an important advancement in producing high yields of fumarate in a sustainable and ecologically-friendly manner.

Opposing effects of bile acids deoxycholic acid and ursodeoxycholic acid on signal transduction pathways in oesophageal cancer cells.

Science.gov (United States)

Abdel-Latif, Mohamed M; Inoue, Hiroyasu; Reynolds, John V

2016-09-01

Ursodeoxycholic acid (UDCA) was reported to reduce bile acid toxicity, but the mechanisms underlying its cytoprotective effects are not fully understood. The aim of the present study was to examine the effects of UDCA on the modulation of deoxycholic acid (DCA)-induced signal transduction in oesophageal cancer cells. Nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) activity was assessed using a gel shift assay. NF-κB activation and translocation was performed using an ELISA-based assay and immunofluorescence analysis. COX-2 expression was analysed by western blotting and COX-2 promoter activity was assessed by luciferase assay. DCA induced NF-κB and AP-1 DNA-binding activities in SKGT-4 and OE33 cells. UDCA pretreatment inhibited DCA-induced NF-κB and AP-1 activation and NF-κB translocation. This inhibitory effect was coupled with a blockade of IκB-α degradation and inhibition of phosphorylation of IKK-α/β and ERK1/2. Moreover, UDCA pretreatment inhibited COX-2 upregulation. Using transient transfection of the COX-2 promoter, UDCA pretreatment abrogated DCA-induced COX-2 promoter activation. In addition, UDCA protected oesophageal cells from the apoptotic effects of deoxycholate. Our findings indicate that UDCA inhibits DCA-induced signalling pathways in oesophageal cancer cells. These data indicate a possible mechanistic role for the chemopreventive actions of UDCA in oesophageal carcinogenesis.

Genetic analysis of pathway regulation for enhancing branched-chain amino acid biosynthesis in plants

KAUST Repository

Chen, Hao

2010-08-01

The branched-chain amino acids (BCAAs) valine, leucine and isoleucine are essential amino acids that play critical roles in animal growth and development. Animals cannot synthesize these amino acids and must obtain them from their diet. Plants are the ultimate source of these essential nutrients, and they synthesize BCAAs through a conserved pathway that is inhibited by its end products. This feedback inhibition has prevented scientists from engineering plants that accumulate high levels of BCAAs by simply over-expressing the respective biosynthetic genes. To identify components critical for this feedback regulation, we performed a genetic screen for Arabidopsis mutants that exhibit enhanced resistance to BCAAs. Multiple dominant allelic mutations in the VALINE-TOLERANT 1 (VAT1) gene were identified that conferred plant resistance to valine inhibition. Map-based cloning revealed that VAT1 encodes a regulatory subunit of acetohydroxy acid synthase (AHAS), the first committed enzyme in the BCAA biosynthesis pathway. The VAT1 gene is highly expressed in young, rapidly growing tissues. When reconstituted with the catalytic subunit in vitro, the vat1 mutant-containing AHAS holoenzyme exhibits increased resistance to valine. Importantly, transgenic plants expressing the mutated vat1 gene exhibit valine tolerance and accumulate higher levels of BCAAs. Our studies not only uncovered regulatory characteristics of plant AHAS, but also identified a method to enhance BCAA accumulation in crop plants that will significantly enhance the nutritional value of food and feed. © 2010 Blackwell Publishing Ltd.

Involvement of salicylic acid, ethylene and jasmonic acid signalling pathways in the susceptibility of tomato to Fusarium oxysporum.

Science.gov (United States)

Di, Xiaotang; Gomila, Jo; Takken, Frank L W

2017-09-01

Phytohormones, such as salicylic acid (SA), ethylene (ET) and jasmonic acid (JA), play key roles in plant defence following pathogen attack. The involvement of these hormones in susceptibility following Fusarium oxysporum (Fo) infection has mostly been studied in Arabidopsis thaliana. However, Fo causes vascular wilt disease in a broad range of crops, including tomato (Solanum lycopersicum). Surprisingly little is known about the involvement of these phytohormones in the susceptibility of tomato towards Fo f. sp. lycopersici (Fol). Here, we investigate their involvement by the analysis of the expression of ET, JA and SA marker genes following Fol infection, and by bioassays of tomato mutants affected in either hormone production or perception. Fol inoculation triggered the expression of SA and ET marker genes, showing the activation of these pathways. NahG tomato, in which SA is degraded, became hypersusceptible to Fol infection and showed stronger disease symptoms than wild-type. In contrast, ACD and Never ripe (Nr) mutants, in which ET biosynthesis and perception, respectively, are impaired, showed decreased disease symptoms and reduced fungal colonization on infection. The susceptibility of the def1 tomato mutant, and a prosystemin over-expressing line, in which JA signalling is compromised or constitutively activated, respectively, was unaltered. Our results show that SA is a negative and ET a positive regulator of Fol susceptibility. The SA and ET signalling pathways appear to act synergistically, as an intact ET pathway is required for the induction of an SA marker gene, and vice versa. © 2017 THE AUTHORS. MOLECULAR PLANT PATHOLOGY PUBLISHED BY BRITISH SOCIETY FOR PLANT PATHOLOGY AND JOHN WILEY & SONS LTD.

Rewiring a secondary metabolite pathway towards itaconic acid production in Aspergillus niger.

Science.gov (United States)

Hossain, Abeer H; Li, An; Brickwedde, Anja; Wilms, Lars; Caspers, Martien; Overkamp, Karin; Punt, Peter J

2016-07-28

The industrially relevant filamentous fungus Aspergillus niger is widely used in industry for its secretion capabilities of enzymes and organic acids. Biotechnologically produced organic acids promise to be an attractive alternative for the chemical industry to replace petrochemicals. Itaconic acid (IA) has been identified as one of the top twelve building block chemicals which have high potential to be produced by biotechnological means. The IA biosynthesis cluster (cadA, mttA and mfsA) has been elucidated in its natural producer Aspergillus terreus and transferred to A. niger to enable IA production. Here we report the rewiring of a secondary metabolite pathway towards further improved IA production through the overexpression of a putative cytosolic citrate synthase citB in a A. niger strain carrying the IA biosynthesis cluster. We have previously shown that expression of cadA from A. terreus results in itaconic acid production in A. niger AB1.13, albeit at low levels. This low-level production is boosted fivefold by the overexpression of mttA and mfsA in itaconic acid producing AB1.13 CAD background strains. Controlled batch cultivations with AB1.13 CAD + MFS + MTT strains showed increased production of itaconic acid compared with AB1.13 CAD strain. Moreover, preliminary RNA-Seq analysis of an itaconic acid producing AB1.13 CAD strain has led to the identification of the putative cytosolic citrate synthase citB which was induced in an IA producing strain. We have overexpressed citB in a AB1.13 CAD + MFS + MTT strain and by doing so hypothesize to have targeted itaconic acid production to the cytosolic compartment. By overexpressing citB in AB1.13 CAD + MFS + MTT strains in controlled batch cultivations we have achieved highly increased titers of up to 26.2 g/L IA with a productivity of 0.35 g/L/h while no CA was produced. Expression of the IA biosynthesis cluster in Aspergillus niger AB1.13 strain enables IA production. Moreover, in the AB1.13 CAD

Terpenoid biosynthesis in Euphorbia lathyris and Copaifera spp

Energy Technology Data Exchange (ETDEWEB)

Skrukrud, C.L.

1987-07-01

Biosynthesis of triterpenoids by isolated latex of Euphorbia lathyris was investigated. The rate of in vitro incorporation of mevalonic acid into triterpenoids was thirty times greater than acetate incorporation indicating that the rate-limiting step in the pathway occurs prior to mevalonate. Both HMG-CoA reductase (EC 1.1.1.34) and HMG-CoA lyase (EC 4.1.3.4) activities were detected in isolated latex. HMG-CoA reductase was localized to a membrane-bound fraction of a 5000g pellet of latex. The rate of conversion of HMG-CoA to mevalonate by this enzyme is comparable to the overall rate of acetate incorporation into the triterpenoids suggesting that this enzyme is rate-determining in the biosynthesis of triterpenoids in E. lathyris latex. HMG-CoA reductase of E. lathyris vegetative tissue was localized to the membrane-bound portion of a particulate fraction (18,000g), and was solubilized by treatment with 2% polyoxyethylene ether W-1. Differences in the optimal pH for activity of HMG-CoA reductase from the latex and vegetative tissue suggest that isozymes of the enzyme may be present in the two tissue types. Studies of the incorporation of various precursors into leaf discs and cuttings taken from Copaifera spp. show differences in the rate of incorporation into Copaifera sesquiterpenes suggesting that the site of sesquiterpene biosynthesis may differ in its accessibility to the different substrates and/or reflecting the metabolic controls on carbon allocation to the terpenes. Mevalonate incorporation by Copaifera langsdorfii cuttings into sesquiterpenes was a hundred-fold greater than either acetate or glucose incorporation, however, its incorporation into squalene and triterpenoids was also a hundred-fold greater than the incorporation into sesquiterpenes. 119 refs., 58 figs., 16 tabs.

Terpenoid biosynthesis in Euphorbia lathyris and Copaifera spp

International Nuclear Information System (INIS)

Skrukrud, C.L.

1987-07-01

Biosynthesis of triterpenoids by isolated latex of Euphorbia lathyris was investigated. The rate of in vitro incorporation of mevalonic acid into triterpenoids was thirty times greater than acetate incorporation indicating that the rate-limiting step in the pathway occurs prior to mevalonate. Both HMG-CoA reductase (EC 1.1.1.34) and HMG-CoA lyase (EC 4.1.3.4) activities were detected in isolated latex. HMG-CoA reductase was localized to a membrane-bound fraction of a 5000g pellet of latex. The rate of conversion of HMG-CoA to mevalonate by this enzyme is comparable to the overall rate of acetate incorporation into the triterpenoids suggesting that this enzyme is rate-determining in the biosynthesis of triterpenoids in E. lathyris latex. HMG-CoA reductase of E. lathyris vegetative tissue was localized to the membrane-bound portion of a particulate fraction (18,000g), and was solubilized by treatment with 2% polyoxyethylene ether W-1. Differences in the optimal pH for activity of HMG-CoA reductase from the latex and vegetative tissue suggest that isozymes of the enzyme may be present in the two tissue types. Studies of the incorporation of various precursors into leaf discs and cuttings taken from Copaifera spp. show differences in the rate of incorporation into Copaifera sesquiterpenes suggesting that the site of sesquiterpene biosynthesis may differ in its accessibility to the different substrates and/or reflecting the metabolic controls on carbon allocation to the terpenes. Mevalonate incorporation by Copaifera langsdorfii cuttings into sesquiterpenes was a hundred-fold greater than either acetate or glucose incorporation, however, its incorporation into squalene and triterpenoids was also a hundred-fold greater than the incorporation into sesquiterpenes. 119 refs., 58 figs., 16 tabs

Oxidation of benzoic acid by heat-activated persulfate: Effect of temperature on transformation pathway and product distribution.

Science.gov (United States)

Zrinyi, Nick; Pham, Anh Le-Tuan

2017-09-01

Heat activates persulfate (S 2 O 8 2- ) into sulfate radical (SO 4 - ), a powerful oxidant capable of transforming a wide variety of contaminants. Previous studies have shown that an increase in temperature accelerates the rates of persulfate activation and contaminant transformation. However, few studies have considered the effect of temperature on contaminant transformation pathway. The objective of this study was to determine how temperature (T = 22-70 °C) influences the activation of persulfate, the transformation of benzoic acid (i.e., a model compound), and the distribution of benzoic acid oxidation products. The time-concentration profiles of the products suggest that benzoic acid was transformed via decarboxylation and hydroxylation mechanisms, with the former becoming increasingly important at elevated temperatures. The pathway through which the products were further oxidized was also influenced by the temperature of persulfate activation. Our findings suggest that the role of temperature in the persulfate-based treatment systems is not limited only to controlling the rates of sulfate and hydroxyl radical generation. The ability of sulfate radical to initiate decarboxylation reactions and, more broadly, fragmentation reactions, as well as the effect of temperature on these transformation pathways could be important to the transformation of a number of organic contaminants. Copyright © 2017 Elsevier Ltd. All rights reserved.

Combinatorial analysis of enzymatic bottlenecks of L-tyrosine pathway by p-coumaric acid production in Saccharomyces cerevisiae.

Science.gov (United States)

Mao, Jiwei; Liu, Quanli; Song, Xiaofei; Wang, Hesuiyuan; Feng, Hui; Xu, Haijin; Qiao, Mingqiang

2017-07-01

To identify new enzymatic bottlenecks of L-tyrosine pathway for further improving the production of L-tyrosine and its derivatives. When ARO4 and ARO7 were deregulated by their feedback resistant derivatives in the host strains, the ARO2 and TYR1 genes, coding for chorismate synthase and prephenate dehydrogenase were further identified as new important rate-limiting steps. The yield of p-coumaric acid in the feedback-resistant strain overexpressing ARO2 or TYR1, was significantly increased from 6.4 to 16.2 and 15.3 mg l -1 , respectively. Subsequently, we improved the strain by combinatorial engineering of pathway genes increasing the yield of p-coumaric acid by 12.5-fold (from 1.7 to 21.3 mg l -1 ) compared with the wild-type strain. Batch cultivations revealed that p-coumaric acid production was correlated with cell growth, and the formation of by-product acetate of the best producer NK-M6 increased to 31.1 mM whereas only 19.1 mM acetate was accumulated by the wild-type strain. Combinatorial metabolic engineering provides a new strategy for further improvement of L-tyrosine or other metabolic biosynthesis pathways in S. cerevisiae.

Carbon isotopic patterns of amino acids associated with various microbial metabolic pathways and physiological conditions

Science.gov (United States)

Wang, P. L.; Hsiao, K. T.; Lin, L. H.

2017-12-01

Amino acids represent one of the most important categories of biomolecule. Their abundance and isotopic patterns have been broadly used to address issues related to biochemical processes and elemental cycling in natural environments. Previous studies have shown that various carbon assimilative pathways of microorganisms (e.g. autotrophy, heterotrophy and acetotrophy) could be distinguished by carbon isotopic patterns of amino acids. However, the taxonomic and catabolic coverage are limited in previous examination. This study aims to uncover the carbon isotopic patterns of amino acids for microorganisms remaining uncharacterized but bearing biogeochemical and ecological significance in anoxic environments. To fulfill the purpose, two anaerobic strains were isolated from riverine wetland and mud volcano in Taiwan. One strain is a sulfate reducing bacterium (related to Desulfovibrio marrakechensis), which is capable of utilizing either H2 or lactate, and the other is a methanogen (related to Methanolobus profundi), which grows solely with methyl-group compounds. Carbon isotope analyses of amino acids were performed on cells grown in exponential and stationary phase. The isotopic patterns were similar for all examined cultures, showing successive 13C depletion along synthetic pathways. No significant difference was observed for the methanogen and lactate-utilizing sulfate reducer harvested in exponential and stationary phases. In contrast, the H2-utilizing sulfate reducer harvested in stationary phase depleted and enriched 13C in aspartic acid and glycine, respectively when compared with that harvested in exponential phase. Such variations might infer the change of carbon flux during synthesis of these two amino acids in the reverse TCA cycle. In addition, the discriminant function analysis for all available data from culture studies further attests the capability of using carbon isotope patterns of amino acids in identifying microbial metabolisms.

Glycyrrhizin, silymarin, and ursodeoxycholic acid regulate a common hepatoprotective pathway in HepG2 cells.

Science.gov (United States)

Hsiang, Chien-Yun; Lin, Li-Jen; Kao, Shung-Te; Lo, Hsin-Yi; Chou, Shun-Ting; Ho, Tin-Yun

2015-07-15

Glycyrrhizin, silymarin, and ursodeoxycholic acid are widely used hepatoprotectants for the treatment of liver disorders, such as hepatitis C virus infection, primary biliary cirrhosis, and hepatocellular carcinoma. The gene expression profiles of HepG2 cells responsive to glycyrrhizin, silymarin, and ursodeoxycholic acid were analyzed in this study. HepG2 cells were treated with 25 µM hepatoprotectants for 24 h. Gene expression profiles of hepatoprotectants-treated cells were analyzed by oligonucleotide microarray in triplicates. Nuclear factor-κB (NF-κB) activities were assessed by luciferase assay. Among a total of 30,968 genes, 252 genes were commonly regulated by glycyrrhizin, silymarin, and ursodeoxycholic acid. These compounds affected the expression of genes relevant various biological pathways, such as neurotransmission, and glucose and lipid metabolism. Genes involved in hepatocarcinogenesis, apoptosis, and anti-oxidative pathways were differentially regulated by all compounds. Moreover, interaction networks showed that NF-κB might play a central role in the regulation of gene expression. Further analysis revealed that these hepatoprotectants inhibited NF-κB activities in a dose-dependent manner. Our data suggested that glycyrrhizin, silymarin, and ursodeoxycholic acid regulated the expression of genes relevant to apoptosis and oxidative stress in HepG2 cells. Moreover, the regulation by these hepatoprotectants might be relevant to the suppression of NF-κB activities. Copyright © 2015 Elsevier GmbH. All rights reserved.

Palmitic acid follows a different metabolic pathway than oleic acid in human skeletal muscle cells; lower lipolysis rate despite an increased level of adipose triglyceride lipase.

Science.gov (United States)

Bakke, Siril S; Moro, Cedric; Nikolić, Nataša; Hessvik, Nina P; Badin, Pierre-Marie; Lauvhaug, Line; Fredriksson, Katarina; Hesselink, Matthijs K C; Boekschoten, Mark V; Kersten, Sander; Gaster, Michael; Thoresen, G Hege; Rustan, Arild C

2012-10-01

Development of insulin resistance is positively associated with dietary saturated fatty acids and negatively associated with monounsaturated fatty acids. To clarify aspects of this difference we have compared the metabolism of oleic (OA, monounsaturated) and palmitic acids (PA, saturated) in human myotubes. Human myotubes were treated with 100μM OA or PA and the metabolism of [(14)C]-labeled fatty acid was studied. We observed that PA had a lower lipolysis rate than OA, despite a more than two-fold higher protein level of adipose triglyceride lipase after 24h incubation with PA. PA was less incorporated into triacylglycerol and more incorporated into phospholipids after 24h. Supporting this, incubation with compounds modifying lipolysis and reesterification pathways suggested a less influenced PA than OA metabolism. In addition, PA showed a lower accumulation than OA, though PA was oxidized to a relatively higher extent than OA. Gene set enrichment analysis revealed that 24h of PA treatment upregulated lipogenesis and fatty acid β-oxidation and downregulated oxidative phosphorylation compared to OA. The differences in lipid accumulation and lipolysis between OA and PA were eliminated in combination with eicosapentaenoic acid (polyunsaturated fatty acid). In conclusion, this study reveals that the two most abundant fatty acids in our diet are partitioned toward different metabolic pathways in muscle cells, and this may be relevant to understand the link between dietary fat and skeletal muscle insulin resistance. Copyright © 2012 Elsevier B.V. All rights reserved.

Bile acid synthesis in man. In vivo activity of the 25-hydroxylation pathway

International Nuclear Information System (INIS)

Duane, W.C.; Pooler, P.A.; Hamilton, J.N.

1988-01-01

During biosynthesis of bile acid, carbons 25-26-27 are removed from the cholesterol side-chain. Side-chain oxidation begins either with hydroxylation at the 26-position, in which case the three-carbon fragment is released as propionic acid, or with hydroxylation at the 25-position, in which case the three-carbon fragment is released as acetone. We have previously shown in the rat that the contribution of the 25-hydroxylation pathway can be quantitated in vivo by measuring production of [ 14 C]acetone from [ 14 C]26-cholesterol. In the present study, we adapted this method to human subjects. 4 d after oral administration of 100 microCi of [ 14 C]26-cholesterol and 1 d after beginning a constant infusion of 16.6 mumol/min unlabeled acetone, three men and two women underwent breath collections. Expired acetone was trapped and purified as the 2,4 dinitrophenylhydrazine derivative. 14 CO 2 was trapped quantitatively using phenethylamine. Specific activity of breath acetone was multiplied by the acetone infusion rate to calculate production of [ 14 C]acetone. [ 14 C]Acetone production averaged 4.9% of total release of 14 C from [ 14 C]26-cholesterol, estimated by 14 CO2 output. The method was validated by showing that [ 14 C]acetone production from [ 14 C]isopropanol averaged 86.9% of the [ 14 C]-isopropanol infusion rate. We conclude that in man, as in the rat, the 25-hydroxylation pathway accounts for less than 5% of bile acid synthesis

A metabolic pathway for catabolizing levulinic acid in bacteria

International Nuclear Information System (INIS)

Rand, Jacqueline M.; Pisithkul, Tippapha; Clark, Ryan L.; Thiede, Joshua M.; Mehrer, Christopher R.

2017-01-01

Microorganisms can catabolize a wide range of organic compounds and therefore have the potential to perform many industrially relevant bioconversions. One barrier to realizing the potential of biorefining strategies lies in our incomplete knowledge of metabolic pathways, including those that can be used to assimilate naturally abundant or easily generated feedstocks. For instance, levulinic acid (LA) is a carbon source that is readily obtainable as a dehydration product of lignocellulosic biomass and can serve as the sole carbon source for some bacteria. Yet, the genetics and structure of LA catabolism have remained unknown. Here, we report the identification and characterization of a seven-gene operon that enables LA catabolism in Pseudomonas putida KT2440. When the pathway was reconstituted with purified proteins, we observed the formation of four acyl-CoA intermediates, including a unique 4-phosphovaleryl-CoA and the previously observed 3-hydroxyvaleryl-CoA product. Using adaptive evolution, we obtained a mutant of Escherichia coli LS5218 with functional deletions of fadE and atoC that was capable of robust growth on LA when it expressed the five enzymes from the P. putida operon. Here, this discovery will enable more efficient use of biomass hydrolysates and metabolic engineering to develop bioconversions using LA as a feedstock.

EPA, DHA, and Lipoic Acid Differentially Modulate the n-3 Fatty Acid Biosynthetic Pathway in Atlantic Salmon Hepatocytes.

Science.gov (United States)

Bou, Marta; Østbye, Tone-Kari; Berge, Gerd M; Ruyter, Bente

2017-03-01

The aim of the present study was to investigate how EPA, DHA, and lipoic acid (LA) influence the different metabolic steps in the n-3 fatty acid (FA) biosynthetic pathway in hepatocytes from Atlantic salmon fed four dietary levels (0, 0.5, 1.0 and 2.0%) of EPA, DHA or a 1:1 mixture of these FA. The hepatocytes were incubated with [1- 14 C] 18:3n-3 in the presence or absence of LA (0.2 mM). Increased endogenous levels of EPA and/or DHA and LA exposure both led to similar responses in cells with reduced desaturation and elongation of [1- 14 C] 18:3n-3 to 18:4n-3, 20:4n-3, and EPA, in agreement with reduced expression of the Δ6 desaturase gene involved in the first step of conversion. DHA production, on the other hand, was maintained even in groups with high endogenous levels of DHA, possibly due to a more complex regulation of this last step in the n-3 metabolic pathway. Inhibition of the Δ6 desaturase pathway led to increased direct elongation to 20:3n-3 by both DHA and LA. Possibly the route by 20:3n-3 and then Δ8 desaturation to 20:4n-3, bypassing the first Δ6 desaturase step, can partly explain the maintained or even increased levels of DHA production. LA increased DHA production in the phospholipid fraction of hepatocytes isolated from fish fed 0 and 0.5% EPA and/or DHA, indicating that LA has the potential to further increase the production of this health-beneficial FA in fish fed diets with low levels of EPA and/or DHA.

Central Nervous System Infection with Borna Disease Virus Causes Kynurenine Pathway Dysregulation and Neurotoxic Quinolinic Acid Production.

Science.gov (United States)

Formisano, Simone; Hornig, Mady; Yaddanapudi, Kavitha; Vasishtha, Mansi; Parsons, Loren H; Briese, Thomas; Lipkin, W Ian; Williams, Brent L

2017-07-15

Central nervous system infection of neonatal and adult rats with Borna disease virus (BDV) results in neuronal destruction and behavioral abnormalities with differential immune-mediated involvement. Neuroactive metabolites generated from the kynurenine pathway of tryptophan degradation have been implicated in several human neurodegenerative disorders. Here, we report that brain expression of key enzymes in the kynurenine pathway are significantly, but differentially, altered in neonatal and adult rats with BDV infection. Gene expression analysis of rat brains following neonatal infection showed increased expression of kynurenine amino transferase II (KATII) and kynurenine-3-monooxygenase (KMO) enzymes. Additionally, indoleamine 2,3-dioxygenase (IDO) expression was only modestly increased in a brain region- and time-dependent manner in neonatally infected rats; however, its expression was highly increased in adult infected rats. The most dramatic impact on gene expression was seen for KMO, whose activity promotes the production of neurotoxic quinolinic acid. KMO expression was persistently elevated in brain regions of both newborn and adult BDV-infected rats, with increases reaching up to 86-fold. KMO protein levels were increased in neonatally infected rats and colocalized with neurons, the primary target cells of BDV infection. Furthermore, quinolinic acid was elevated in neonatally infected rat brains. We further demonstrate increased expression of KATII and KMO, but not IDO, in vitro in BDV-infected C6 astroglioma cells. Our results suggest that BDV directly impacts the kynurenine pathway, an effect that may be exacerbated by inflammatory responses in immunocompetent hosts. Thus, experimental models of BDV infection may provide new tools for discriminating virus-mediated from immune-mediated impacts on the kynurenine pathway and their relative contribution to neurodegeneration. IMPORTANCE BDV causes persistent, noncytopathic infection in vitro yet still elicits

Electron microscopy of hydrocarbon production in parthenium argentatum (guayule)

Energy Technology Data Exchange (ETDEWEB)

Bauer, Thomas E. [Univ. of California, Berkeley, CA (United States)

1977-11-01

The electron microscope was used to study the biological processes involved in hydrocarbon production. The little desert shrub Guayule (Parthenium argentatum) was selected for study. This shrub can produce hydrocarbons (rubber) in concentrations up to 1/4 of its dry weight. It grows on semi-arid land and has been extensively studied. The potential of Guayule is described in detail. Results of an investigation into the morphology of Guayule at the electron microscope level are given. Experiments, which would allow the biosynthesis of hydrocarbon in Guayule to be followed, were designed. In order to do this, knowledge of the biochemistry of rubber formation was used to select a tracer, mevalonic acid. Mevalonic acid is the precursor of all the terpenoids, a large class of hydrocarbons which includes rubber. It was found that when high enough concentrations of mevalonic acid are administered to seedling Guayule plants, build-ups of metabolized products are found within the chloroplasts of the seedlings. Also, tritium labeled mevalonic acid was used as a precursor, and its metabolic progress was followed by using the technique of electron microscope autoradiography. The results of these experiments also implicated chloroplasts of the Guayule plant in hydrocarbon production. The final task was the development of a system to produce three-dimensional stereo reconstructions of organelles suspected of involvement in hydrocarbon biosynthesis in Guayule. The techniques are designed to reconstruct an object from serial sections of that object. The techniques use stereo imaging both to abstract information for computer processing, and also in the computer produced reconstruction.

Indole-3-acetic acid biosynthetic pathway and aromatic amino acid aminotransferase activities in Pantoea dispersa strain GPK.

Science.gov (United States)

Kulkarni, G B; Nayak, A S; Sajjan, S S; Oblesha, A; Karegoudar, T B

2013-05-01

This investigation deals with the production of IAA by a bacterial isolate Pantoea dispersa strain GPK (PDG) identified by 16S rRNA gene sequence analysis. HPLC and Mass spectral analysis of metabolites from bacterial spent medium revealed that, IAA production by PDG is Trp-dependent and follows indole-3-pyruvic acid (IPyA) pathway. Substrate specificity study of aromatic amino acid aminotransferase (AAT) showed high activities, only when tryptophan (Trp) and α-ketoglutarate (α-kg) were used as substrates. AAT is highly specific for Trp and α-kg as amino group donor and acceptor, respectively. The effect of exogenous IAA on bacterial growth was established. Low concentration of exogenous IAA induced the growth, whereas high concentration decreased the growth of bacterium. PDG treatment significantly increased the root length, shoot length and dry mass of the chickpea and pigeon pea plants. © 2013 The Society for Applied Microbiology.

Compound-Specific Carbon, Nitrogen, and Hydrogen Isotopic Ratios for Amino Acids in CM and CR Chondrites and their use in Evaluating Potential Formation Pathways

Science.gov (United States)

Elsila, Jamie E.; Charnley, Steven B.; Burton, Aaron S.; Glavin, Daniel P.; Dworkin, Jason P.

2012-01-01

Stable hydrogen, carbon, and nitrogen isotopic ratios (oD, 013C, and olSN) of organic compounds can revcal information about their origin and formation pathways. Several formation mechanisms and environments have been postulated for the amino acids detected in carbonaceous chondrites. As each proposed mechanism utilizes different precursor molecules, the isotopic signatures of the resulting amino acids may indicate the most likely of these pathways. We have applied gas chromatography with mass spectrometry and combustion isotope ratio mass spectrometry to measure the compound-specific C, N, and H stable isotopic ratios of amino acids from seven CM and CR carbonaceous chondrites: CM1I2 Allan Hills (ALH) 83100, CM2 Murchison, CM2 Lewis Cliff (LEW) 90500, CM2 Lonewolf Nunataks (LON) 94101, CRZ Graves Nunataks (GRA) 95229, CRZ Elephant Moraine (EET) 92042, and CR3 Queen Alexandra Range (QUE) 99177. We compare the isotopic compositions of amino acids in these meteorites with predictions of expected isotopic enrichments from potential formation pathways. We observe trends of decreasing ODC and increasing oD with increasing carbon number in the aH, (l-NH2 amino acids that correspond to predictions made for formation via Streckercyanohydrin synthesis. We also observe light ODC signatures for -alanine, which may indicate either formation via Michael addition or via a pathway that forms primarily small, straight-chain, amine-terminal amino acids (n-ro-amino acids). Higher deuterium enrichments are observed in amethyl amino acids, indicating formation of these amino acids or their precursors in cold interstellar or nebular environments. Finally, individual amino acids are more enriched in deuterium in CR chondrites than CM chondrites, reflecting different parent-body chemistry.

Imaging Preferential Flow Pathways of Contaminants from Passive Acid Mine Drainage Mitigation Sites Using Electrical Resistivity

Science.gov (United States)

Kelley, N.; Mount, G.; Terry, N.; Herndon, E.; Singer, D. M.

2017-12-01

The Critical Zone represents the surficial and shallow layer of rock, air, water, and soil where most interactions between living organisms and the Earth occur. Acid mine drainage (AMD) resulting from coal extraction can influence both biological and geochemical processes across this zone. Conservative estimates suggest that more than 300 million gallons of AMD are released daily, making this acidic solution of water and contaminants a common issue in areas with legacy or current coal extraction. Electrical resistivity imaging (ERI) provides a rapid and minimally invasive method to identify and monitor contaminant pathways from AMD remediation systems in the subsurface of the Critical Zone. The technique yields spatially continuous data of subsurface resistivity that can be inverted to determine electrical conductivity as a function of depth. Since elevated concentrations of heavy metals can directly influence soil conductivity, ERI data can be used to trace the flow pathways or perhaps unknown mine conduits and transport of heavy metals through the subsurface near acid mine drainage sources. This study aims to examine preferential contaminant migration from those sources through substrate pores, fractures, and shallow mine workings in the near subsurface surrounding AMD sites in eastern Ohio and western Pennsylvania. We utilize time lapse ERI measures during different hydrologic conditions to better understand the variability of preferential flow pathways in relation to changes in stage and discharge within the remediation systems. To confirm ERI findings, and provide constraint to geochemical reactions occurring in the shallow subsurface, we conducted Inductively Coupled Plasma (ICP) spectrometry analysis of groundwater samples from boreholes along the survey transects. Through these combined methods, we can provide insight into the ability of engineered systems to contain and isolate metals in passive acid mine drainage treatment systems.

Degradation of clofibric acid in UV/chlorine disinfection process: kinetics, reactive species contribution and pathways.

Science.gov (United States)

Tang, Yuqing; Shi, Xueting; Liu, Yongze; Feng, Li; Zhang, Liqiu

2018-02-01

As a potential endocrine disruptor, clofibric acid (CA) was investigated in this study for its degradation kinetics and pathways in UV/chlorine process. The results showed that CA in both UV photolysis and UV/chlorine processes could be degraded via pseudo-first-order kinetics, while it almost could not be degraded in the dark chlorination process. The observed rate constant ( k obs ) in UV photolysis was 0.0078 min -1, and increased to 0.0107 min -1 combining with 0.1 mM chlorine. The k obs increased to 0.0447 min -1 with further increasing the chlorine dosage from 0.1 to 1.0 mM, and reached a plateau at higher dosage (greater than 1.0 mM). The higher k obs was obtained at acid solution rather than basic solution. Moreover, the calculated contributions of radical species to k obs indicated that the HO• contributed significantly to CA degradation in acidic conditions, while the reactive chlorine species and UV direct photolysis dominated in neutral and basic solution. The degradation of CA was slightly inhibited in the presence of [Formula: see text] (1 ∼ 50 mM), barely affected by the presence of Cl - (1 ∼ 200 mM) and greatly suppressed by humic acid (0 ∼ 5 mg l -1 ). Thirteen main degradation intermediates and three degradation pathways of CA were identified during UV/chlorine process.

Biosynthetic pathways to delta-aminolevulinic acid induced by blue light in the pigment mutant C-2A' of Scenedesmus obliquus

International Nuclear Information System (INIS)

Klein, O.; Senger, H.

1978-01-01

The X-ray induced mutant C-2A' of Scenedesmus obliquus grows heterotrophically but forms only traces of chlorophyll in the dark. Upon illumination, delta-aminolevulinic acid (ALA) is synthesized and chlorophyll is formed. These processes are blue light dependent and ceased immediately when the cells were transferred back into darkness. Addition of levulinic acid (LA) inhibited the light-dependent formation of chlorophyll and caused accumulation of ALA by competitive inhibition of the ALA dehydratase (EC. 4.2.1.24). By feeding specifically labelled 14 C precursors to the pigment mutant, inhibiting the ALA dehydratase with LA, accumulating, extracting and analyzing the ALA, two pathways leading towards ALA could be established: glycine and succinyl CoA can be condensed to ALA and the 5 carbon skeleton of glutamate can completely be incorporated into ALA via a second pathway. The glycine-succinyl CoA pathway dominated over the glutamate pathway, but both led to chlorophyll formation. (author)

Acetic acid activates the AMP-activated protein kinase signaling pathway to regulate lipid metabolism in bovine hepatocytes.

Directory of Open Access Journals (Sweden)

Xinwei Li

Full Text Available The effect of acetic acid on hepatic lipid metabolism in ruminants differs significantly from that in monogastric animals. Therefore, the aim of this study was to investigate the regulation mechanism of acetic acid on the hepatic lipid metabolism in dairy cows. The AMP-activated protein kinase (AMPK signaling pathway plays a key role in regulating hepatic lipid metabolism. In vitro, bovine hepatocytes were cultured and treated with different concentrations of sodium acetate (neutralized acetic acid and BML-275 (an AMPKα inhibitor. Acetic acid consumed a large amount of ATP, resulting in an increase in AMPKα phosphorylation. The increase in AMPKα phosphorylation increased the expression and transcriptional activity of peroxisome proliferator-activated receptor α, which upregulated the expression of lipid oxidation genes, thereby increasing lipid oxidation in bovine hepatocytes. Furthermore, elevated AMPKα phosphorylation reduced the expression and transcriptional activity of the sterol regulatory element-binding protein 1c and the carbohydrate responsive element-binding protein, which reduced the expression of lipogenic genes, thereby decreasing lipid biosynthesis in bovine hepatocytes. In addition, activated AMPKα inhibited the activity of acetyl-CoA carboxylase. Consequently, the triglyceride content in the acetate-treated hepatocytes was significantly decreased. These results indicate that acetic acid activates the AMPKα signaling pathway to increase lipid oxidation and decrease lipid synthesis in bovine hepatocytes, thereby reducing liver fat accumulation in dairy cows.

Metabolic engineering of Escherichia coli for limonene and perillyl alcohol production.

Science.gov (United States)

Alonso-Gutierrez, Jorge; Chan, Rossana; Batth, Tanveer S; Adams, Paul D; Keasling, Jay D; Petzold, Christopher J; Lee, Taek Soon

2013-09-01

Limonene is a valuable monoterpene used in the production of several commodity chemicals and medicinal compounds. Among them, perillyl alcohol (POH) is a promising anti-cancer agent that can be produced by hydroxylation of limonene. We engineered E. coli with a heterologous mevalonate pathway and limonene synthase for production of limonene followed by coupling with a cytochrome P450, which specifically hydroxylates limonene to produce POH. A strain containing all mevalonate pathway genes in a single plasmid produced limonene at titers over 400mg/L from glucose, substantially higher than has been achieved in the past. Incorporation of a cytochrome P450 to hydroxylate limonene yielded approximately 100mg/L of POH. Further metabolic engineering of the pathway and in situ product recovery using anion exchange resins would make this engineered E. coli a potential production platform for any valuable limonene derivative. © 2013 Elsevier Inc. All rights reserved.

De novo transcriptome assembly and the putative biosynthetic pathway of steroidal sapogenins of Dioscorea composita.

Directory of Open Access Journals (Sweden)

Xia Wang

Full Text Available The plant Dioscorea composita has important applications in the medical and energy industries, and can be used for the extraction of steroidal sapogenins (important raw materials for the synthesis of steroidal drugs and bioethanol production. However, little is known at the genetic level about how sapogenins are biosynthesized in this plant. Using Illumina deep sequencing, 62,341 unigenes were obtained by assembling its transcriptome, and 27,720 unigenes were annotated. Of these, 8,022 unigenes were mapped to 243 specific pathways, and 531 unigenes were identified to be involved in 24 secondary metabolic pathways. 35 enzymes, which were encoded by 79 unigenes, were related to the biosynthesis of steroidal sapogenins in this transcriptome database, covering almost all the nodes in the steroidal pathway. The results of real-time PCR experiments on ten related transcripts (HMGR, MK, SQLE, FPPS, DXS, CAS, HMED, CYP51, DHCR7, and DHCR24 indicated that sapogenins were mainly biosynthesized by the mevalonate pathway. The expression of these ten transcripts in the tuber and leaves was found to be much higher than in the stem. Also, expression in the shoots was low. The nucleotide and protein sequences and conserved domains of four related genes (HMGR, CAS, SQS, and SMT1 were highly conserved between D. composita and D. zingiberensis; but expression of these four genes is greater in D. composita. However, there is no expression of these key enzymes in potato and no steroidal sapogenins are synthesized.

Regulation of the Omega-3 Fatty Acid Biosynthetic Pathway in Atlantic Salmon Hepatocytes.

Directory of Open Access Journals (Sweden)

Marte Avranden Kjær

Full Text Available Limited availability of the n-3 fatty acids EPA and DHA have led to an interest in better understanding of the n-3 biosynthetic pathway and its regulation. The biosynthesis of alpha-linolenic acid to EPA and DHA involves several complex reaction steps including desaturation-, elongation- and peroxisomal beta-oxidation enzymes. The aims of the present experiments were to gain more knowledge on how this biosynthesis is regulated over time by different doses and fatty acid combinations. Hepatocytes isolated from salmon were incubated with various levels and combinations of oleic acid, EPA and DHA. Oleic acid led to a higher expression of the Δ6 fatty acid desaturase (fad genes Δ6fad_a, Δ6fad_b, Δ6fad_c and the elongase genes elovl2 compared with cells cultured in medium enriched with DHA. Further, the study showed rhythmic variations in expression over time. Levels were reached where a further increase in specific fatty acids given to the cells not stimulated the conversion further. The gene expression of Δ6fad_a_and Δ6fad_b responded similar to fatty acid treatment, suggesting a co-regulation of these genes, whereas Δ5fad and Δ6fad_c showed a different regulation pattern. EPA and DHA induced different gene expression patterns, especially of Δ6fad_a. Addition of radiolabelled alpha-linolenic acid to the hepatocytes confirmed a higher degree of elongation and desaturation in cells treated with oleic acid compared to cells treated with DHA. This study suggests a complex regulation of the conversion process of n-3 fatty acids. Several factors, such as that the various gene copies are differently regulated, the gene expression show rhythmic variations and gene expression only affected to a certain level, determines when you get the maximum conversion of the beneficial n-3 fatty acids.

Quercetin, ascorbic acid, caffeine and ellagic acid are more efficient than rosiglitazone, metformin and glimepiride in interfering with pathways leading to the development of neurological complications associated with diabetes: A comparative in-vitro study

Directory of Open Access Journals (Sweden)

Vineet Mehta

2017-06-01

Full Text Available Neuropathy is the least understood and most devastating complication associated with diabetes. Diabetic neuropathy develops in patients despite of regular therapy, indicating that marketed drugs has minimal effect on pathways leading to the development and progression of these complications. Present study was aimed to evaluate natural compounds for their ability to interfere with pathways leading to the development of diabetes mediated neurological complications and compare their efficacy with marketed anti-diabetic drugs. Anti-diabetic potential of ascorbic acid, gallic acid, quercetin, ellagic acid, cinnamic acid, caffeine and piperine was predicted by evaluating in-silico interaction energy (kcal/mol of these compounds with insulin receptor, peroxisome proliferator-activated receptor gamma-γ and dipeptidyl peptidase-4 proteins. Ascorbic acid, gallic acid, quercetin and ellagic acid showed excellent in-vitro antioxidant activity in DPPH radical scavenging and inhibition of lipid peroxidation assay, which was 1.5–3 folds better than the marketed drugs. Quercetin, gallic acid, cinnamic acid, piperine and caffeine efficiently prevented H2O2 induced genotoxicity, which commercial drugs failed to prevent. Further, quercetin, ellagic acid, caffeine and ascorbic acid were 3–4.7 folds better than marketed drugs in inhibiting α-amylase activity. Herbal molecules and rosiglitazone showed comparable results for glucose uptake, which may be attributed to enhanced GLUT4 translocation into primary neuronal culture under hyperglycemic conditions. In conclusion, currently available marketed anti-diabetic drugs have minimal effect on the pathways leading to diabetic neuropathy and supplementing diabetic therapeutics with quercetin, ascorbic acid, caffeine and ellagic acid may be better suited to counter diabetic neuropathy through inhibiting oxidative stress, genotoxicity and improving neuronal glucose utilization.

The mitochondrial fatty acid synthesis (mtFASII) pathway is capable of mediating nuclear-mitochondrial cross talk through the PPAR system of transcriptional activation

International Nuclear Information System (INIS)

Parl, Angelika; Mitchell, Sabrina L.; Clay, Hayley B.; Reiss, Sara; Li, Zhen; Murdock, Deborah G.

2013-01-01

Highlights: •The function of the mitochondria fatty acid synthesis pathway is partially unknown. •Overexpression of the pathway causes transcriptional activation through PPARs. •Knock down of the pathway attenuates that activation. •The last enzyme in the pathway regulates its own transcription. •Products of the mtFASII pathway are able to drive nuclear transcription. -- Abstract: Mammalian cells contain two fatty acid synthesis pathways, the cytosolic FASI pathway, and the mitochondrial FASII pathway. The selection behind the conservation of the mitochondrial pathway is not completely understood, given the presence of the cytosolic FAS pathway. In this study, we show through heterologous gene reporter systems and PCR-based arrays that overexpression of MECR, the last step in the mtFASII pathway, causes modulation of gene expression through the PPAR pathway. Electromobility shift assays (EMSAs) demonstrate that overexpression of MECR causes increased binding of PPARs to DNA, while cell fractionation and imaging studies show that MECR remains localized to the mitochondria. Interestingly, knock down of the mtFASII pathway lessens the effect of MECR on this transcriptional modulation. Our data are most consistent with MECR-mediated transcriptional activation through products of the mtFASII pathway, although we cannot rule out MECR acting as a coactivator. Further investigation into the physiological relevance of this communication will be necessary to better understand some of the phenotypic consequences of deficits in this pathway observed in animal models and human disease

The mitochondrial fatty acid synthesis (mtFASII) pathway is capable of mediating nuclear-mitochondrial cross talk through the PPAR system of transcriptional activation

Energy Technology Data Exchange (ETDEWEB)

Parl, Angelika; Mitchell, Sabrina L.; Clay, Hayley B.; Reiss, Sara; Li, Zhen; Murdock, Deborah G., E-mail: deborah.murdock@vanderbilt.edu

2013-11-15

Highlights: •The function of the mitochondria fatty acid synthesis pathway is partially unknown. •Overexpression of the pathway causes transcriptional activation through PPARs. •Knock down of the pathway attenuates that activation. •The last enzyme in the pathway regulates its own transcription. •Products of the mtFASII pathway are able to drive nuclear transcription. -- Abstract: Mammalian cells contain two fatty acid synthesis pathways, the cytosolic FASI pathway, and the mitochondrial FASII pathway. The selection behind the conservation of the mitochondrial pathway is not completely understood, given the presence of the cytosolic FAS pathway. In this study, we show through heterologous gene reporter systems and PCR-based arrays that overexpression of MECR, the last step in the mtFASII pathway, causes modulation of gene expression through the PPAR pathway. Electromobility shift assays (EMSAs) demonstrate that overexpression of MECR causes increased binding of PPARs to DNA, while cell fractionation and imaging studies show that MECR remains localized to the mitochondria. Interestingly, knock down of the mtFASII pathway lessens the effect of MECR on this transcriptional modulation. Our data are most consistent with MECR-mediated transcriptional activation through products of the mtFASII pathway, although we cannot rule out MECR acting as a coactivator. Further investigation into the physiological relevance of this communication will be necessary to better understand some of the phenotypic consequences of deficits in this pathway observed in animal models and human disease.

The effect of MEP pathway and other inhibitors on the intracellular localization of a plasma membrane-targeted, isoprenylable GFP reporter protein in tobacco BY-2 cells

Science.gov (United States)

Bach, Thomas J

2013-01-01

We have established an in vivo visualization system for the geranylgeranylation of proteins in a stably transformed tobacco BY-2 cell line, based on the expression of a dexamethasone-inducible GFP fused to the carboxy-terminal basic domain of the rice calmodulin CaM61, which naturally bears a CaaL geranylgeranylation motif (GFP-BD-CVIL). By using pathway-specific inhibitors it was demonstrated that inhibition of the methylerythritol phosphate (MEP) pathway with known inhibitors like oxoclomazone and fosmidomycin, as well as inhibition of the protein geranylgeranyltransferase type 1 (PGGT-1), shifted the localization of the GFP-BD-CVIL protein from the membrane to the nucleus. In contrast, the inhibition of the mevalonate (MVA) pathway with mevinolin did not affect the localization. During the present work, this test system has been used to examine the effect of newly designed inhibitors of the MEP pathway and inhibitors of sterol biosynthesis such as squalestatin, terbinafine and Ro48-8071. In addition, we also studied the impact of different post-prenylation inhibitors or those suspected to affect the transport of proteins to the plasma membrane on the localization of the geranylgeranylable fusion protein GFP-BD-CVIL. PMID:24555083

Coregulation of terpenoid pathway genes and prediction of isoprene production in Bacillus subtilis using transcriptomics

Energy Technology Data Exchange (ETDEWEB)

Hess, Becky M.; Xue, Junfeng; Markillie, Lye Meng; Taylor, Ronald C.; Wiley, H. S.; Ahring, Birgitte K.; Linggi, Bryan E.

2013-06-19

The isoprenoid pathway converts pyruvate to isoprene and related isoprenoid compounds in plants and some bacteria. Currently, this pathway is of great interest because of the critical role that isoprenoids play in basic cellular processes as well as the industrial value of metabolites such as isoprene. Although the regulation of several pathway genes has been described, there is a paucity of information regarding the system level regulation and control of the pathway. To address this limitation, we examined Bacillus subtilis grown under multiple conditions and then determined the relationship between altered isoprene production and the pattern of gene expression. We found that terpenoid genes appeared to fall into two distinct subsets with opposing correlations with respect to the amount of isoprene produced. The group whose expression levels positively correlated with isoprene production included dxs, the gene responsible for the commitment step in the pathway, as well as ispD, and two genes that participate in the mevalonate pathway, yhfS and pksG. The subset of terpenoid genes that inversely correlated with isoprene production included ispH, ispF, hepS, uppS, ispE, and dxr. A genome wide partial least squares regression model was created to identify other genes or pathways that contribute to isoprene production. This analysis showed that a subset of 213 regulated genes was sufficient to create a predictive model of isoprene production under different conditions and showed correlations at the transcriptional level. We conclude that gene expression levels alone are sufficiently informative about the metabolic state of a cell that produces increased isoprene and can be used to build a model which accurately predicts production of this secondary metabolite across many simulated environmental conditions.

Poly(lactic-co-glycolic) acid drug delivery systems through transdermal pathway: an overview

OpenAIRE

Naves, Lucas; Dhand, Chetna; Almeida, Luis; Rajamani, Lakshminarayanan; Ramakrishna, Seeram; Soares, Gra?a

2017-01-01

In past few decades, scientists have made tremendous advancement in the field of drug delivery systems (DDS), through transdermal pathway, as the skin represents a ready and large surface area for delivering drugs. Efforts are in progress to design efficient transdermal DDS that support sustained drug release at the targeted area for longer duration in the recommended therapeutic window without producing side-effects. Poly(lactic-co-glycolic acid) (PLGA) is one of the most promising Food and ...

Two zebrafish G2A homologs activate multiple intracellular signaling pathways in acidic environment

Energy Technology Data Exchange (ETDEWEB)

Ichijo, Yuta; Mochimaru, Yuta [Laboratory of Cell Signaling Regulation, Department of Life Sciences, School of Agriculture, Meiji University, Kawasaki 214-8571 (Japan); Azuma, Morio [Laboratory of Regulatory Biology, Graduate School of Science and Engineering, University of Toyama, 3190-Gofuku, Toyama 930-8555 (Japan); Satou, Kazuhiro; Negishi, Jun [Laboratory of Cell Signaling Regulation, Department of Life Sciences, School of Agriculture, Meiji University, Kawasaki 214-8571 (Japan); Nakakura, Takashi [Department of Anatomy, Graduate School of Medicine, Teikyo University, 2-11-1 Itabashi-Ku, Tokyo 173-8605 (Japan); Oshima, Natsuki [Laboratory of Cell Signaling Regulation, Department of Life Sciences, School of Agriculture, Meiji University, Kawasaki 214-8571 (Japan); Mogi, Chihiro; Sato, Koichi [Laboratory of Signal Transduction, Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512 (Japan); Matsuda, Kouhei [Laboratory of Regulatory Biology, Graduate School of Science and Engineering, University of Toyama, 3190-Gofuku, Toyama 930-8555 (Japan); Okajima, Fumikazu [Laboratory of Signal Transduction, Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512 (Japan); Tomura, Hideaki, E-mail: tomurah@meiji.ac.jp [Laboratory of Cell Signaling Regulation, Department of Life Sciences, School of Agriculture, Meiji University, Kawasaki 214-8571 (Japan)

2016-01-01

Human G2A is activated by various stimuli such as lysophosphatidylcholine (LPC), 9-hydroxyoctadecadienoic acid (9-HODE), and protons. The receptor is coupled to multiple intracellular signaling pathways, including the G{sub s}-protein/cAMP/CRE, G{sub 12/13}-protein/Rho/SRE, and G{sub q}-protein/phospholipase C/NFAT pathways. In the present study, we examined whether zebrafish G2A homologs (zG2A-a and zG2A-b) could respond to these stimuli and activate multiple intracellular signaling pathways. We also examined whether histidine residue and basic amino acid residue in the N-terminus of the homologs also play roles similar to those played by human G2A residues if the homologs sense protons. We found that the zG2A-a showed the high CRE, SRE, and NFAT activities, however, zG2A-b showed only the high SRE activity under a pH of 8.0. Extracellular acidification from pH 7.4 to 6.3 ameliorated these activities in zG2A-a-expressing cells. On the other hand, acidification ameliorated the SRE activity but not the CRE and NFAT activities in zG2A-b-expressing cells. LPC or 9-HODE did not modify any activity of either homolog. The substitution of histidine residue at the 174{sup th} position from the N-terminus of zG2A-a to asparagine residue attenuated proton-induced CRE and NFAT activities but not SRE activity. The substitution of arginine residue at the 32nd position from the N-terminus of zG2A-a to the alanine residue also attenuated its high and the proton-induced CRE and NFAT activities. On the contrary, the substitution did not attenuate SRE activity. The substitution of the arginine residue at the 10th position from the N-terminus of zG2A-b to the alanine residue also did not attenuate its high or the proton-induced SRE activity. These results indicate that zebrafish G2A homologs were activated by protons but not by LPC and 9-HODE, and the activation mechanisms of the homologs were similar to those of human G2A. - Highlights: • Zebrafish two G2A homologs are proton

8,9-Epoxyeicosatrienoic acid analog protects pulmonary artery smooth muscle cells from apoptosis via ROCK pathway

International Nuclear Information System (INIS)

Ma, Jun; Zhang, Lei; Li, Shanshan; Liu, Shulin; Ma, Cui; Li, Weiyang; Falck, J.R.; Manthati, Vijay L.; Reddy, D. Sudarshan; Medhora, Meetha; Jacobs, Elizabeth R.; Zhu, Daling

2010-01-01

Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid (AA) catalyzed by cytochrome P450 (CYP), have many essential biologic roles in the cardiovascular system including inhibition of apoptosis in cardiomyocytes. In the present study, we tested the potential of 8,9-EET and derivatives to protect pulmonary artery smooth muscle cells (PASMCs) from starvation induced apoptosis. We found 8,9-epoxy-eicos-11(Z)-enoic acid (8,9-EET analog (214)), but not 8,9-EET, increased cell viability, decreased activation of caspase-3 and caspase-9, and decreased TUNEL-positive cells or nuclear condensation induced by serum deprivation (SD) in PASMCs. These effects were reversed after blocking the Rho-kinase (ROCK) pathway with Y-27632 or HA-1077. Therefore, 8,9-EET analog (214) protects PASMC from serum deprivation-induced apoptosis, mediated at least in part via the ROCK pathway. Serum deprivation of PASMCs resulted in mitochondrial membrane depolarization, decreased expression of Bcl-2 and enhanced expression of Bax, all effects were reversed by 8,9-EET analog (214) in a ROCK dependent manner. Because 8,9-EET and not the 8,9-EET analog (214) protects pulmonary artery endothelial cells (PAECs), these observations suggest the potential to differentially promote apoptosis or survival with 8,9-EET or analogs in pulmonary arteries.

8,9-Epoxyeicosatrienoic acid analog protects pulmonary artery smooth muscle cells from apoptosis via ROCK pathway

Energy Technology Data Exchange (ETDEWEB)

Ma, Jun; Zhang, Lei; Li, Shanshan [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Liu, Shulin [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Bio-pharmaceutical Key Laboratory of Heilongjiang Province, Harbin 150081 (China); Ma, Cui [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Li, Weiyang [Mudanjiang Medical College, Mudanjiang 157011 (China); Falck, J.R.; Manthati, Vijay L.; Reddy, D. Sudarshan [University of Texas Southwestern Medical Center, Dallas, TX 75390 (United States); Medhora, Meetha; Jacobs, Elizabeth R. [Division of Pulmonary and Critical Care, Department of Medicine, Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Zhu, Daling, E-mail: dalingz@yahoo.com [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Bio-pharmaceutical Key Laboratory of Heilongjiang Province, Harbin 150081 (China)

2010-08-15

Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid (AA) catalyzed by cytochrome P450 (CYP), have many essential biologic roles in the cardiovascular system including inhibition of apoptosis in cardiomyocytes. In the present study, we tested the potential of 8,9-EET and derivatives to protect pulmonary artery smooth muscle cells (PASMCs) from starvation induced apoptosis. We found 8,9-epoxy-eicos-11(Z)-enoic acid (8,9-EET analog (214)), but not 8,9-EET, increased cell viability, decreased activation of caspase-3 and caspase-9, and decreased TUNEL-positive cells or nuclear condensation induced by serum deprivation (SD) in PASMCs. These effects were reversed after blocking the Rho-kinase (ROCK) pathway with Y-27632 or HA-1077. Therefore, 8,9-EET analog (214) protects PASMC from serum deprivation-induced apoptosis, mediated at least in part via the ROCK pathway. Serum deprivation of PASMCs resulted in mitochondrial membrane depolarization, decreased expression of Bcl-2 and enhanced expression of Bax, all effects were reversed by 8,9-EET analog (214) in a ROCK dependent manner. Because 8,9-EET and not the 8,9-EET analog (214) protects pulmonary artery endothelial cells (PAECs), these observations suggest the potential to differentially promote apoptosis or survival with 8,9-EET or analogs in pulmonary arteries.

Activation of type 2 cannabinoid receptors (CB2R) promotes fatty acid oxidation through the SIRT1/PGC-1α pathway

Energy Technology Data Exchange (ETDEWEB)

Zheng, Xuqin [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China); Sun, Tao [Department of Neurology, Jinling Hospital, Nanjing University School of Medicine, Nanjing, Jiangsu Province 210002 (China); Wang, Xiaodong, E-mail: xdwang666@hotmail.com [Department of Endocrinology, First Affiliated Hospital, Nanjing Medical University, Nanjing, Jiangsu Province 210029 (China)

2013-07-05

Highlights: •TC, a CB2R specific agonist, stimulates SIRT1 activity by PKA/CREB pathway. •TC promotes PGC-1α transcriptional activity by increasing its deacetylation. •TC increases the expression of genes linked to FAO and promotes the rate of FAO. •The effects of TC in FAO are dependent on CB2R. •Suggesting CB2R as a target to treat diseases with lipid dysregulation. -- Abstract: Abnormal fatty acid oxidation has been associated with obesity and type 2 diabetes. At the transcriptional level, peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) has been reported to strongly increase the ability of hormone nuclear receptors PPARα and ERRα to drive transcription of fatty acid oxidation enzymes. In this study, we report that a specific agonist of the type 2 cannabinoid receptor (CB2R) can lead to fatty acid oxidation through the PGC-1α pathway. We have found that CB2R is expressed in differentiated C2C12 myotubes, and that use of the specific agonist trans-caryophyllene (TC) stimulates sirtuin 1 (SIRT1) deacetylase activity by increasing the phosphorylation of cAMP response element-binding protein (CREB), thus leading to increased levels of PGC-1α deacetylation. This use of TC treatment increases the expression of genes linked to the fatty acid oxidation pathway in a SIRT1/PGC-1α-dependent mechanism and also drastically accelerates the rate of complete fatty acid oxidation in C2C12 myotubes, neither of which occur when CB2R mRNA is knocked down using siRNA. These results reveal that activation of CB2R by a selective agonist promotes lipid oxidation through a signaling/transcriptional pathway. Our findings imply that pharmacological manipulation of CB2R may provide therapeutic possibilities to treat metabolic diseases associated with lipid dysregulation.

Gallic acid modulates phenotypic behavior and gene expression in oral squamous cell carcinoma cells by interfering with leptin pathway.

Science.gov (United States)

Santos, Eliane Macedo Sobrinho; da Rocha, Rogério Gonçalves; Santos, Hércules Otacílio; Guimarães, Talita Antunes; de Carvalho Fraga, Carlos Alberto; da Silveira, Luiz Henrique; Batista, Paulo Ricardo; de Oliveira, Paulo Sérgio Lopes; Melo, Geraldo Aclécio; Santos, Sérgio Henrique; de Paula, Alfredo Maurício Batista; Guimarães, André Luiz Sena; Farias, Lucyana Conceição

2018-01-01

Gallic acid is a polyphenolic compost appointed to interfere with neoplastic cells behavior. Evidence suggests an important role of leptin in carcinogenesis pathways, inducing a proliferative phenotype. We investigated the potential of gallic acid to modulate leptin-induced cell proliferation and migration of oral squamous cell carcinoma cell lines. The gallic acid effect on leptin secretion by oral squamous cell carcinoma cells, as well as the underlying molecular mechanisms, was also assessed. For this, we performed proliferation, migration, immunocytochemical and qPCR assays. The expression levels of cell migration-related genes (MMP2, MMP9, Col1A1, and E-cadherin), angiogenesis (HIF-1α, mir210), leptin signaling (LepR, p44/42 MAPK), apoptosis (casp-3), and secreted leptin levels by oral squamous cell carcinoma cells were also measured. Gallic acid decreased proliferation and migration of leptin-treated oral squamous cell carcinoma cells, and reduced mRNA expression of MMP2, MMP9, Col1A1, mir210, but did not change HIF-1α. Gallic acid decreased levels of leptin secreted by oral squamous cell carcinoma cells, accordingly with downregulation of p44/42 MAPK expression. Thus, gallic acid appears to break down neoplastic phenotype of oral squamous cell carcinoma cells by interfering with leptin pathway. Copyright © 2017 Elsevier GmbH. All rights reserved.

Acidic preparations of lysed platelets upregulate proliferative pathways in osteoblast-like cells as demonstrated by genome-wide microarray analysis.

Science.gov (United States)

Wahlström, Ola; Linder, Cecilia Halling; Ansell, Anna; Kalén, Anders; Söderström, Mats; Magnusson, Per

2011-01-01

Platelets contain numerous growth factors essential for wound and fracture healing. We investigated the gene expression in human osteoblast-like cells stimulated with lysed platelets prepared in acidic, neutral, or alkaline buffers. Lysed platelets prepared in buffers at pH 5.4, 7.4, and 7.9, were added after neutralization to hFOB 1.19 cells. Genome-wide microarray analysis was performed using the Affymetrix GeneChip 7G Scanner. Biometric, cluster, and pathway analyses were performed with GeneSpring GX. Biometric analyses demonstrated that 53 genes were differentially regulated (p ≤ 0.005, ≥2-fold increase). Pathway analysis revealed 10 significant pathways of which eight are common ones regulating bone formation and cancer growth. Eleven genes were selected for quantitative real-time polymerase chain reaction (PCR) based on the microarray analysis of the lysed platelets prepared in the pH 5.4 experiments. In conclusion, acidic preparations of lysed platelet concentrates release factors essential for cell proliferation and particularly cell metabolism under hypoxic conditions. The genetic response from these factors was dominated by genes associated with the same pathways observed in bone formation and cancer growth. Activation of TGF-β in the acidic preparation could be a stimulatory key factor of cell proliferation. These results support the hypothesis that acidification of platelets modifies the stimulatory response of mesenchymal cells in vitro, which is analogous with the observed milieu of a low pH present in wound and fracture sites, as well as in growing tumors.

Rational engineering of p-hydroxybenzoate hydroxylase to enable efficient gallic acid synthesis via a novel artificial biosynthetic pathway.

Science.gov (United States)

Chen, Zhenya; Shen, Xiaolin; Wang, Jian; Wang, Jia; Yuan, Qipeng; Yan, Yajun

2017-11-01

Gallic acid (GA) is a naturally occurring phytochemical that has strong antioxidant and antibacterial activities. It is also used as a potential platform chemical for the synthesis of diverse high-value compounds. Hydrolytic degradation of tannins by acids, bases or microorganisms serves as a major way for GA production, which however, might cause environmental pollution and low yield and efficiency. Here, we report a novel approach for efficient microbial production of GA. First, structure-based rational engineering of PobA, a p-hydroxybenzoate hydroxylase from Pseudomonas aeruginosa, generated a new mutant, Y385F/T294A PobA, which displayed much higher activity toward 3,4-dihydroxybenzoic acid (3,4-DHBA) than the wild-type and any other reported mutants. Remarkably, expression of this mutant in Escherichia coli enabled generation of 1149.59 mg/L GA from 1000 mg/L 4-hydroxybenzoic acid (4-HBA), representing a 93% molar conversion ratio. Based on that, we designed and reconstituted a novel artificial biosynthetic pathway of GA and achieved 440.53 mg/L GA production from simple carbon sources in E. coli. Further enhancement of precursor supply through reinforcing shikimate pathway was able to improve GA de novo production to 1266.39 mg/L in shake flasks. Overall, this study not only led to the development of a highly active PobA variant for hydroxylating 3,4-DHBA into GA via structure-based protein engineering approach, but also demonstrated a promising pathway for bio-based manufacturing of GA and its derived compounds. Biotechnol. Bioeng. 2017;114: 2571-2580. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana

Energy Technology Data Exchange (ETDEWEB)

Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China); Bian, Po, E-mail: bianpo@ipp.ac.cn [Key Laboratory of Ion Beam Bioengineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, Hefei 230031 (China); Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei 230031 (China)

2016-09-15

Highlights: • The JA signal pathway plays a pivotal role in mediating radiation-induced bystander effects in Arabidopsis thaliana. • The JA signal pathway is involved in both the generation of bystander signals in irradiated roots and radiation responses in bystander aerial plants. • Over-accumulation of endogenous JA enhances the radiosensitivity of plants in terms of RIBE. - Abstract: Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX

A pivotal role of the jasmonic acid signal pathway in mediating radiation-induced bystander effects in Arabidopsis thaliana.

Science.gov (United States)

Wang, Ting; Xu, Wei; Deng, Chenguang; Xu, Shaoxin; Li, Fanghua; Wu, Yuejin; Wu, Lijun; Bian, Po

Although radiation-induced bystander effects (RIBE) in Arabidopsis thaliana have been well demonstrated in vivo, little is known about their underlying mechanisms, particularly with regard to the participating signaling molecules and signaling pathways. In higher plants, jasmonic acid (JA) and its bioactive derivatives are well accepted as systemic signal transducers that are produced in response to various environmental stresses. It is therefore speculated that the JA signal pathway might play a potential role in mediating radiation-induced bystander signaling of root-to-shoot. In the present study, pretreatment of seedlings with Salicylhydroxamic acid, an inhibitor of lipoxigenase (LOX) in JA biosynthesis, significantly suppressed RIBE-mediated expression of the AtRAD54 gene. After root irradiation, the aerial parts of A. thaliana mutants deficient in JA biosynthesis (aos) and signaling cascades (jar1-1) showed suppressed induction of the AtRAD54 and AtRAD51 genes and TSI and 180-bp repeats, which have been extensively used as endpoints of bystander genetic and epigenetic effects in plants. These results suggest an involvement of the JA signal pathway in the RIBE of plants. Using the root micro-grafting technique, the JA signal pathway was shown to participate in both the generation of bystander signals in irradiated root cells and radiation responses in the bystander aerial parts of plants. The over-accumulation of endogenous JA in mutant fatty acid oxygenation up-regulated 2 (fou2), in which mutation of the Two Pore Channel 1 (TPC1) gene up-regulates expression of the LOX and allene oxide synthase (AOS) genes, inhibited RIBE-mediated expression of the AtRAD54 gene, but up-regulated expression of the AtKU70 and AtLIG4 genes in the non-homologous end joining (NHEJ) pathway. Considering that NHEJ is employed by plants with increased DNA damage, the switch from HR to NHEJ suggests that over-accumulation of endogenous JA might enhance the radiosensitivity of plants

Effect of uric acid on inflammatory COX-2 and ROS pathways in vascular smooth muscle cells.

Science.gov (United States)

Oğuz, Nurgül; Kırça, Mustafa; Çetin, Arzu; Yeşilkaya, Akın

2017-10-01

Hyperuricemia is thought to play a role in cardiovascular diseases (CVD), including hypertension, coronary artery disease and atherosclerosis. However, exactly how uric acid contributes to these pathologies is unknown. An underlying mechanism of inflammatory diseases, such as atherosclerosis, includes enhanced production of cyclooxygenase-2 (COX-2) and superoxide anion. Here, we aimed to examine the effect of uric acid on inflammatory COX-2 and superoxide anion production and to determine the role of losartan. Primarily cultured vascular smooth muscle cells (VSMCs) were time and dose-dependently induced by uric acid and COX-2 and superoxide anion levels were measured. COX-2 levels were determined by ELISA, and superoxide anion was measured by the superoxide dismutase (SOD)-inhibitable reduction of ferricytochrome c method. Uric acid elevated COX-2 levels in a time-dependent manner. Angiotensin-II receptor blocker, losartan, diminished uric-acid-induced COX-2 elevation. Uric acid also increased superoxide anion level in VSMCs. Uric acid plays an important role in CVD pathogenesis by inducing inflammatory COX-2 and ROS pathways. This is the first study demonstrating losartan's ability to reduce uric-acid-induced COX-2 elevation.

An overview of the non-mevalonate pathway for terpenoid

Indian Academy of Sciences (India)

Unknown

troversial role of isomerase via non-MVA route in which both IPP and DMAPP are reported to be synthe- ... chemical scheme was proposed with a head-to-head con- densation of ..... berry exocarp and mesocarp; Phytochemistry 60 451–459.

Statins inhibit protein lipidation and induce the unfolded protein response in the non-sterol producing nematode Caenorhabditis elegans

DEFF Research Database (Denmark)

Mörck, Catarina; Elmelund-Præstekær, Louise Cathrine Braun; Kurth, Caroline

2009-01-01

of lipid moieties for protein prenylation. The nematode Caenorhabditis elegans possesses a mevalonate pathway that lacks the branch leading to cholesterol synthesis, and thus represents an ideal organism to specifically study the noncholesterol roles of the pathway. Inhibiting HMG-CoA reductase in C....... elegans using statins or RNAi leads to developmental arrest and loss of membrane association of a GFP-based prenylation reporter. The unfolded protein response (UPR) is also strongly activated, suggesting that impaired prenylation of small GTPases leads to the accumulation of unfolded proteins and ER...... and fatty acid composition were unaffected in statin-treated worms, even though they showed reduced staining with Nile red. We conclude that inhibitors of HMG-CoA reductase or of farnesyl transferases induce the UPR by inhibiting the prenylation of M57.2 substrates, resulting in developmental arrest in C...

Gallic acid inhibits vascular calcification through the blockade of BMP2-Smad1/5/8 signaling pathway.

Science.gov (United States)

Kee, Hae Jin; Cho, Soo-Na; Kim, Gwi Ran; Choi, Sin Young; Ryu, Yuhee; Kim, In Kyeom; Hong, Young Joon; Park, Hyung Wook; Ahn, Youngkeun; Cho, Jeong Gwan; Park, Jong Chun; Jeong, Myung Ho

2014-11-01

Vascular calcification is associated with increased risk of morbidity and mortality in patients with cardiovascular diseases, chronic kidney diseases, and diabetes. Gallic acid, a natural compound found in gallnut and green tea, is known to be antifungal, antioxidant, and anticancer. Here we investigated the effect of gallic acid on vascular smooth muscle cell (VSMC) calcification and the underlying mechanism. Gallic acid inhibited inorganic phosphate-induced osteoblast differentiation markers as well as calcification phenotypes (as determined by calcium deposition, Alizarin Red, and Von Kossa staining). Knockdown of BMP2 or Noggin blocked phosphate-induced calcification. Gallic acid suppressed phosphorylation of Smad1/5/8 protein induced by inorganic phosphate. Taken together, we suggest that gallic acid acts as a novel therapeutic agent of vascular calcification by mediating BMP2-Smad1/5/8 signaling pathway. Copyright © 2014 Elsevier Inc. All rights reserved.

Meta-analysis of global transcriptomics reveals conserved genetic pathways of Quercetin and Tannic acid mediated longevity in C. elegans

Directory of Open Access Journals (Sweden)

Kerstin ePietsch

2012-04-01

Full Text Available Recent research has highlighted that the polyphenols Quercetin and Tannic acid are capable of extending the lifespan of C. elegans. To gain a deep understanding of the underlying molecular genetics, we analyzed the global transcriptional patterns of nematodes exposed to Quercetin or Tannic acid concentrations that are non-effective (in lifespan extension, lifespan extending or toxic. By means of an intricate meta-analysis it was possible to compare the transcriptomes of polyphenol exposure to recently published data sets derived from i longevity mutants or ii infection. This detailed comparative in silico analysis facilitated the identification of compound specific and overlapping transcriptional profiles and allowed the formulation of mechanistic models of Quercetin and Tannic acid mediated longevity. Lifespan extension due to Quercetin was predominantly driven by the metabolome, TGF-beta signaling, Insulin-like signaling and the p38 MAPK pathway and Tannic acid’s impact involved, in part, the amino acid metabolism and was modulated by the TGF-beta and the p38 MAPK pathways. DAF-12, which integrates TGF-beta and Insulin-like downstream signaling, therefore seems to be a crucial regulator for both polyphenols.

Interactions between the jasmonic and salicylic acid pathway modulate the plant metabolome and affect herbivores of different feeding types.

Science.gov (United States)

Schweiger, R; Heise, A-M; Persicke, M; Müller, C

2014-07-01

The phytohormones jasmonic acid (JA) and salicylic acid (SA) mediate induced plant defences and the corresponding pathways interact in a complex manner as has been shown on the transcript and proteine level. Downstream, metabolic changes are important for plant-herbivore interactions. This study investigated metabolic changes in leaf tissue and phloem exudates of Plantago lanceolata after single and combined JA and SA applications as well as consequences on chewing-biting (Heliothis virescens) and piercing-sucking (Myzus persicae) herbivores. Targeted metabolite profiling and untargeted metabolic fingerprinting uncovered different categories of plant metabolites, which were influenced in a specific manner, indicating points of divergence, convergence, positive crosstalk and pronounced mutual antagonism between the signaling pathways. Phytohormone-specific decreases of primary metabolite pool sizes in the phloem exudates may indicate shifts in sink-source relations, resource allocation, nutrient uptake or photosynthesis. Survival of both herbivore species was significantly reduced by JA and SA treatments. However, the combined application of JA and SA attenuated the negative effects at least against H. virescens suggesting that mutual antagonism between the JA and SA pathway may be responsible. Pathway interactions provide a great regulatory potential for the plant that allows triggering of appropriate defences when attacked by different antagonist species. © 2013 John Wiley & Sons Ltd.

Over-expression of VvWRKY1 in grapevines induces expression of jasmonic acid pathway-related genes and confers higher tolerance to the downy mildew.

Directory of Open Access Journals (Sweden)

Chloé Marchive

Full Text Available Most WRKY transcription factors activate expression of defence genes in a salicylic acid- and/or jasmonic acid-dependent signalling pathway. We previously identified a WRKY gene, VvWRKY1, which is able to enhance tolerance to fungal pathogens when it is overexpressed in tobacco. The present work analyzes the effects of VvWRKY1 overexpression in grapevine. Microarray analysis showed that genes encoding defence-related proteins were up-regulated in the leaves of transgenic 35S::VvWRKY1 grapevines. Quantitative RT-PCR analysis confirmed that three genes putatively involved in jasmonic acid signalling pathway were overexpressed in the transgenic grapes. The ability of VvWRKY1 to trans-activate the promoters of these genes was demonstrated by transient expression in grape protoplasts. The resistance to the causal agent of downy mildew, Plasmopara viticola, was enhanced in the transgenic plants. These results show that VvWRKY1 can increase resistance of grapevine against the downy mildew through transcriptional reprogramming leading to activation of the jasmonic acid signalling pathway.

Statins and protein prenylation in cancer cell biology and therapy.

Science.gov (United States)

Garcia-Ruiz, Carmen; Morales, Albert; Fernandez-Checa, Jose C

2012-05-01

The use of statins has scaled up to become one of the most prescribed medicines in the world and have been very useful in the manegement of cardiovascular diseases and related mortality. The disclosure of their chemical structure similar to that of hydroxy methyl glutaryl-CoA (HMG-CoA) revealed their ability to compete with and inhibit the rate-limiting enzyme HMG-CoA reductase that catalyzes the synthesis of mevalonate, which then serves as the precursor for isoprenoids and cholesterol in the mevalonate pathway. While most of the effects of statins are associated with the lowering of cellular cholesterol levels, it is clear that they also blunt the non-sterol branch of the mevalonate pathway, decreasing formation of isoprenoids and altering protein-prenylation, a critical event in the posttranslational modulation of proteins involved in the regulation of cell cycle progression, proliferation and signaling pathways. Randomized controlled trials for the prevention of cardiovascular diseases indicated that statins elicited provocative and unexpected benefits for reducing a number of different types of cancers, including colorectal carcinoma, melanoma, prostate and hepatocellular carcinoma, although in other cancer types the preclinical expectations of statins were dissapointing. In this review, we will describe the evidence and mechanisms underlying the potential beneficial use of statins and the role of protein prenylation in cancer prevention. Of relevance, the combination of statins with other anti cancer drugs may be a significant asset in malignancies resistant to current therapy.

Characterization of the complete uric acid degradation pathway in the fungal pathogen Cryptococcus neoformans.

Directory of Open Access Journals (Sweden)

I Russel Lee

Full Text Available Degradation of purines to uric acid is generally conserved among organisms, however, the end product of uric acid degradation varies from species to species depending on the presence of active catabolic enzymes. In humans, most higher primates and birds, the urate oxidase gene is non-functional and hence uric acid is not further broken down. Uric acid in human blood plasma serves as an antioxidant and an immune enhancer; conversely, excessive amounts cause the common affliction gout. In contrast, uric acid is completely degraded to ammonia in most fungi. Currently, relatively little is known about uric acid catabolism in the fungal pathogen Cryptococcus neoformans even though this yeast is commonly isolated from uric acid-rich pigeon guano. In addition, uric acid utilization enhances the production of the cryptococcal virulence factors capsule and urease, and may potentially modulate the host immune response during infection. Based on these important observations, we employed both Agrobacterium-mediated insertional mutagenesis and bioinformatics to predict all the uric acid catabolic enzyme-encoding genes in the H99 genome. The candidate C. neoformans uric acid catabolic genes identified were named: URO1 (urate oxidase, URO2 (HIU hydrolase, URO3 (OHCU decarboxylase, DAL1 (allantoinase, DAL2,3,3 (allantoicase-ureidoglycolate hydrolase fusion protein, and URE1 (urease. All six ORFs were then deleted via homologous recombination; assaying of the deletion mutants' ability to assimilate uric acid and its pathway intermediates as the sole nitrogen source validated their enzymatic functions. While Uro1, Uro2, Uro3, Dal1 and Dal2,3,3 were demonstrated to be dispensable for virulence, the significance of using a modified animal model system of cryptococcosis for improved mimicking of human pathogenicity is discussed.

Molecular docking and NMR binding studies to identify novel inhibitors of human phosphomevalonate kinase

Energy Technology Data Exchange (ETDEWEB)

Boonsri, Pornthip [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States); Department of Chemistry, NANOTEC Center of Nanotechnology, National Nanotechnology Center, Faculty of Science, Kasetsart University, Bangkok 10900 (Thailand); Neumann, Terrence S.; Olson, Andrew L.; Cai, Sheng [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States); Herdendorf, Timothy J.; Miziorko, Henry M. [Division of Molecular Biology and Biochemistry, School of Biological Sciences, University of Missouri-Kansas City, Kansas City, MO 64110 (United States); Hannongbua, Supa [Department of Chemistry, NANOTEC Center of Nanotechnology, National Nanotechnology Center, Faculty of Science, Kasetsart University, Bangkok 10900 (Thailand); Sem, Daniel S., E-mail: daniel.sem@cuw.edu [Chemical Proteomics Facility at Marquette, Department of Chemistry, Marquette University, Milwaukee, WI 53201 (United States)

2013-01-04

Highlights: Black-Right-Pointing-Pointer Natural and synthetic inhibitors of human phosphomevalonate kinase identified. Black-Right-Pointing-Pointer Virtual screening yielded a hit rate of 15%, with inhibitor K{sub d}'s of 10-60 {mu}M. Black-Right-Pointing-Pointer NMR studies indicate significant protein conformational changes upon binding. -- Abstract: Phosphomevalonate kinase (PMK) phosphorylates mevalonate-5-phosphate (M5P) in the mevalonate pathway, which is the sole source of isoprenoids and steroids in humans. We have identified new PMK inhibitors with virtual screening, using autodock. Promising hits were verified and their affinity measured using NMR-based {sup 1}H-{sup 15}N heteronuclear single quantum coherence (HSQC) chemical shift perturbation and fluorescence titrations. Chemical shift changes were monitored, plotted, and fitted to obtain dissociation constants (K{sub d}). Tight binding compounds with K{sub d}'s ranging from 6-60 {mu}M were identified. These compounds tended to have significant polarity and negative charge, similar to the natural substrates (M5P and ATP). HSQC cross peak changes suggest that binding induces a global conformational change, such as domain closure. Compounds identified in this study serve as chemical genetic probes of human PMK, to explore pharmacology of the mevalonate pathway, as well as starting points for further drug development.

[Ursodeoxycholic acid induced apoptosis of human hepatoma cells HepG2 and SMMC-7721 bymitochondrial-mediated pathway].

Science.gov (United States)

Wu, Duan; Zhou, Jianyin; Yin, Zhenyu; Liu, Pingguo; Zhao, Yilin; Liu, Jianming; Wang, Xiaomin

2014-12-02

To explore the effects and underlying mechanisms of ursodeoxycholic acid on human hepatoma cells. HepG2 and SMMC-7721 HCC cell lines were respectively treated with ursodeoxycholic acid. And cell proliferation, apoptosis and the expression of Bax/Bcl-2 gene were detected by methyl thiazolyl tetrazolium (MTT), inverted microscopy, fluorescent microscopy, flow cytometry and Western blot. Ursodeoxycholic acid significantly inhibited the proliferation of human hepatoma cells in a concentration- and time-dependent manner. The half maximal inhibitory concentrations (IC50) of HepG2 and SMMC-7721 were 397.3 and 387.7 µg/ml respectively after a 48-hour treatment of 400 µg /ml ursodeoxycholic acid. And it also induced the apoptosis of HepG2 and SMMC-7721 cells, up-regulated Bax gene and down-regulated Bcl-2 gene. Ursodeoxycholic acid inhibits the proliferation of hepatoma cells and induce apoptosis by mitochondrial-mediated pathway.

Evidence for an excitatory amino acid pathway in the brainstem and for its involvement in cardiovascular control.

Science.gov (United States)

Somogyi, P; Minson, J B; Morilak, D; Llewellyn-Smith, I; McIlhinney, J R; Chalmers, J

1989-09-04

The source and possible role of excitatory amino acid projections to areas of the ventrolateral medulla (VLM) involved in cardiovascular control were studied. Following the injection of [3H]D-aspartate ([3H]D-Asp), a selective tracer for excitatory amino acid pathways, into vasopressor or vasodepressor areas of the VLM in rats, more than 90% of retrogradely labelled neurones were found in the nucleus of the solitary tract (NTS). Very few of the [3H]D-Asp-labelled cells were immunoreactive for tyrosine hydroxylase, none for phenylethanolamine-N-methyltransferase or gamma-aminobutyric acid. The density of labelled cells in the NTS was similar to that obtained with the non-selective tracers wheat germ agglutinin-horseradish peroxidase (WGA-HRP) and WGA-colloidal gold, but these tracers also labelled other cell groups in the medulla. Furthermore, the decrease in blood pressure, caused by pharmacological activation of neurones in the NTS of rats, or by electrical stimulation of the aortic depressor nerve in rabbits could be blocked by the selective N-methyl-D-aspartate (NMDA) receptor antagonist 2-amino-5-phosphonovalerate injected into the caudal vasodepressor area of the VLM. This area corresponds to the termination of [3H]D-Asp transporting NTS neurones. These results provide evidence that a population of NTS neurones projecting to the VLM use excitatory amino acids as transmitters. Among other possible functions, this pathway may mediate tonic and reflex control of blood pressure via NMDA receptors in the VLM.

Both the Jasmonic Acid and the Salicylic Acid Pathways Contribute to Resistance to the Biotrophic Clubroot Agent Plasmodiophora brassicae in Arabidopsis.

Science.gov (United States)

Lemarié, Séverine; Robert-Seilaniantz, Alexandre; Lariagon, Christine; Lemoine, Jocelyne; Marnet, Nathalie; Jubault, Mélanie; Manzanares-Dauleux, Maria J; Gravot, Antoine

2015-11-01

The role of salicylic acid (SA) and jasmonic acid (JA) signaling in resistance to root pathogens has been poorly documented. We assessed the contribution of SA and JA to basal and partial resistance of Arabidopsis to the biotrophic clubroot agent Plasmodiophora brassicae. SA and JA levels as well as the expression of the SA-responsive genes PR2 and PR5 and the JA-responsive genes ARGAH2 and THI2.1 were monitored in infected roots of the accessions Col-0 (susceptible) and Bur-0 (partially resistant). SA signaling was activated in Bur-0 but not in Col-0. The JA pathway was weakly activated in Bur-0 but was strongly induced in Col-0. The contribution of both pathways to clubroot resistance was then assessed using exogenous phytohormone application and mutants affected in SA or JA signaling. Exogenous SA treatment decreased clubroot symptoms in the two Arabidopsis accessions, whereas JA treatment reduced clubroot symptoms only in Col-0. The cpr5-2 mutant, in which SA responses are constitutively induced, was more resistant to clubroot than the corresponding wild type, and the JA signaling-deficient mutant jar1 was more susceptible. Finally, we showed that the JA-mediated induction of NATA1 drove N(δ)-acetylornithine biosynthesis in infected Col-0 roots. The 35S::NATA1 and nata1 lines displayed reduced or enhanced clubroot symptoms, respectively, thus suggesting that in Col-0 this pathway was involved in the JA-mediated basal clubroot resistance. Overall, our data support the idea that, depending on the Arabidopsis accession, both SA and JA signaling can play a role in partial inhibition of clubroot development in compatible interactions with P. brassicae. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Artemisinic acid exhibits antitumor activity in MCF-7 breast cancer cells through the inhibition of angiogenesis, VEGF, m-TOR and AKT signalling pathways

Directory of Open Access Journals (Sweden)

Yan Cui

2016-09-01

Full Text Available The aim of the present study was to evaluate the antitumor and anti-angiogenic effects of artemisinic acid in MCF-7 human breast cancer cells. Various cell signalling pathways (VEGF, m-TOR and AKT signalling pathways and MTT assay were used. The in vivo antitumor activity of artemisinic acid was evaluated by means of tumor xenograft mouse model. Transwell cell migration assay was used to examine the chemotactic motility of the human umbilical vascular endothelial cells (HUVECs, while as endothelial cell capillary-like tube formation assay was used to evaluate the effect of artemisinic acid on the tube formation in HUVECs. We found that artemisinic acid considerably reduced both the volume and weight of concrete tumors and reduced angiogenesis in a xenograft mouse tumor model in vivo. Further, artemisinic acid suppressed the VEGF-induced cell migration and capillary-like tube formation of HUVECs in a dose-dependent manner. Artemisinic acid was found to suppress the VEGF-induced phosphorylation of VEGFR2 and also the activity of AKT and m-TOR.

Foeniculum vulgare essential oil ameliorates acetic acid-induced colitis in rats through the inhibition of NF-kB pathway.

Science.gov (United States)

Rezayat, Seyed Mahdi; Dehpour, Ahmad-Reza; Motamed, Saeed Mohammadi; Yazdanparast, Maryam; Chamanara, Mohsen; Sahebgharani, Mousa; Rashidian, Amir

2017-10-24

The aim of the present study is to investigate the protective effects of Foeniculum vulgare essential oil on intestinal inflammation through the inhibition of NF-kB pathway in acetic acid-induced rat colitis. Acute colitis was induced by intra-rectal administration of 2 mL of diluted acetic acid (4%) solution. Two hours after the induction of colitis, 0.2% tween 80 in normal saline, dexamethasone (2 mg/kg) and F. vulgare essential oil (100, 200, 400 mg/kg) were administered to the animals by oral gavage and continued for 5 consecutive days. Assessment of macroscopic and microscopic lesions was done. MPO activity was evaluated by biochemical method. Furthermore, TNF-α activity was detected by immunohistochemistry (IHC) and the expression level of p-NF-kB p65 protein was measured by western blot analysis. Dexamethasone and F. vulgare essential oil (200, 400 mg/kg) reduced the macroscopic and microscopic lesions compared to the acetic acid group (p kB p65 protein (p kB pathway.

Palmitic acid suppresses apolipoprotein M gene expression via the pathway of PPAR{sub β/δ} in HepG2 cells

Energy Technology Data Exchange (ETDEWEB)

Luo, Guanghua; Shi, Yuanping; Zhang, Jun; Mu, Qinfeng; Qin, Li; Zheng, Lu; Feng, Yuehua [Comprehensive Laboratory, The Third Affiliated Hospital of Soochow University, Changzhou 213003 (China); Berggren-Söderlund, Maria; Nilsson-Ehle, Peter [Division of Clinical Chemistry and Pharmacology, Department of Laboratory Medicine, Lund University, S-221 85 Lund (Sweden); Zhang, Xiaoying, E-mail: zhangxy6689996@163.com [Department of Cardiothoracic Surgery, The Third Affiliated Hospital of Soochow University, Changzhou 213003 (China); Xu, Ning, E-mail: ning.xu@med.lu.se [Division of Clinical Chemistry and Pharmacology, Department of Laboratory Medicine, Lund University, S-221 85 Lund (Sweden)

2014-02-28

Highlights: • Palmitic acid significantly inhibited APOM gene expression in HepG2 cells. • Palmitic acid could obviously increase PPARB/D mRNA levels in HepG2 cells. • PPAR{sub β/δ} antagonist, GSK3787, had no effect on APOM expression. • GSK3787 could reverse the palmitic acid-induced down-regulation of APOM expression. • Palmitic acid induced suppression of APOM expression is mediated via the PPAR{sub β/δ} pathway. - Abstract: It has been demonstrated that apolipoprotein M (APOM) is a vasculoprotective constituent of high density lipoprotein (HDL), which could be related to the anti-atherosclerotic property of HDL. Investigation of regulation of APOM expression is of important for further exploring its pathophysiological function in vivo. Our previous studies indicated that expression of APOM could be regulated by platelet activating factor (PAF), transforming growth factors (TGF), insulin-like growth factor (IGF), leptin, hyperglycemia and etc., in vivo and/or in vitro. In the present study, we demonstrated that palmitic acid could significantly inhibit APOM gene expression in HepG2 cells. Further study indicated neither PI-3 kinase (PI3K) inhibitor LY294002 nor protein kinase C (PKC) inhibitor GFX could abolish palmitic acid induced down-regulation of APOM expression. In contrast, the peroxisome proliferator-activated receptor beta/delta (PPAR{sub β/δ}) antagonist GSK3787 could totally reverse the palmitic acid-induced down-regulation of APOM expression, which clearly demonstrates that down-regulation of APOM expression induced by palmitic acid is mediated via the PPAR{sub β/δ} pathway.

The plastidial 2-C-methyl-D-erythritol 4-phosphate pathway provides the isoprenyl moiety for protein geranylgeranylation in tobacco BY-2 cells.

Science.gov (United States)

Gerber, Esther; Hemmerlin, Andréa; Hartmann, Michael; Heintz, Dimitri; Hartmann, Marie-Andrée; Mutterer, Jérôme; Rodríguez-Concepción, Manuel; Boronat, Albert; Van Dorsselaer, Alain; Rohmer, Michel; Crowell, Dring N; Bach, Thomas J

2009-01-01

Protein farnesylation and geranylgeranylation are important posttranslational modifications in eukaryotic cells. We visualized in transformed Nicotiana tabacum Bright Yellow-2 (BY-2) cells the geranylgeranylation and plasma membrane localization of GFP-BD-CVIL, which consists of green fluorescent protein (GFP) fused to the C-terminal polybasic domain (BD) and CVIL isoprenylation motif from the Oryza sativa calmodulin, CaM61. Treatment with fosmidomycin (Fos) or oxoclomazone (OC), inhibitors of the plastidial 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, caused mislocalization of the protein to the nucleus, whereas treatment with mevinolin, an inhibitor of the cytosolic mevalonate pathway, did not. The nuclear localization of GFP-BD-CVIL in the presence of MEP pathway inhibitors was completely reversed by all-trans-geranylgeraniol (GGol). Furthermore, 1-deoxy-d-xylulose (DX) reversed the effects of OC, but not Fos, consistent with the hypothesis that OC blocks 1-deoxy-d-xylulose 5-phosphate synthesis, whereas Fos inhibits its conversion to 2-C-methyl-d-erythritol 4-phosphate. By contrast, GGol and DX did not rescue the nuclear mislocalization of GFP-BD-CVIL in the presence of a protein geranylgeranyltransferase type 1 inhibitor. Thus, the MEP pathway has an essential role in geranylgeranyl diphosphate (GGPP) biosynthesis and protein geranylgeranylation in BY-2 cells. GFP-BD-CVIL is a versatile tool for identifying pharmaceuticals and herbicides that interfere either with GGPP biosynthesis or with protein geranylgeranylation.

Study on the secondary metabolic regulation of callus of Panax quinquefolium L

International Nuclear Information System (INIS)

Zhang Meiping; Wang Yi; Sun Chunyu; Li Xianggao

2003-01-01

The effects of precursors with different kind, concentration, growth phase, time and different ATP, nicotinic acid concentration on the callus growth and saponin formation with the buds of Panax quinquefolium L. has been studied. The results showed that 0.1% mevalonic acid inhibited the callus growth but promoted the saponin formation; L-leucine of 100 mg/L had no obvious effects on callus growth, but promoted the saponin formation; Mg(Ac) 2 of 444 mg/L promoted both the callus growth and saponin content. The optimum time for precursor feeding is at the beginning of regeneration. The optimum time for mevalonic acid is 6h; ATP of 160 mg/L promoted both the callus growth and saponin formation. Nicotinic acid of 1.0 mg/L promoted callus growth and saponin formation. With 4000 Gy of 60 Co γ-rays, the saponin content has been increased 1 time compared with nonirradiation

Multiple signal transduction pathways in okadaic acid induced apoptosis in HeLa cells

International Nuclear Information System (INIS)

Jayaraj, R.; Gupta, Nimesh; Rao, P.V. Lakshmana

2009-01-01

Okadaic acid (OA) is the major component of diarrhetic shell fish poisoning toxins and a potent inhibitor of protein phosphatase 1 and 2A. We investigated the signal transduction pathways involved in OA induced cell death in HeLa cells. OA induced cytotoxicity and apoptosis at IC50 of 100 nM. OA treatment resulted in time dependent increase in reactive oxygen species and depleted intracellular glutathione levels. Loss of mitochondrial membrane permeability led to translocation of bax, cytochrome-c and AIF from mitochondria to cytosol. The cells under fluorescence microscope showed typical apoptotic morphology with condensed chromatin, and nuclear fragmentation. We investigated the mitochondrial-mediated caspase cascade. The time dependent activation and cleavage of of bax, caspases-8, 10, 9, 3 and 7 was observed in Western blot analysis. In addition to caspase-dependent pathway AIF mediated caspase-independent pathway was involved in OA mediated cell death. OA also caused time dependent inhibition of protein phosphatase 2A activity and phosphorylation of p38 and p42/44 MAP kinases. Inhibitor studies with Ac-DEVO-CHO and Z-VAD-FMK could not prevent the phosphorylation of p38 and p42/44 MAP kinases. Our experiments with caspase inhibitors Ac-DEVD-CHO, Z-IETD-FMK and Z-VAD-FMK inhibited capsase-3, 8 cleavages but did not prevent OA-induced apoptosis and DNA fragmentation. Similarly, pretreatment with cyclosporin-A and N-acetylcysteine could not prevent the DNA fragmentation. In summary, the results of our study show that OA induces multiple signal transduction pathways acting either independently or simultaneously leading to apoptosis

Structure of active IspH enzyme from escherichia coli provides mechanistic insights into substrate reduction

KAUST Repository

Grä wert, Tobias; Rohdich, Felix; Span, Lngrid; Backer, Adelbert; Eisenreich, Wolfgang; Eppinger, Jö rg; Groll, Michael

2009-01-01

The terminal step of the non-mevalonate pathway of terpene biosynthesis is catalyzed by IspH (see scheme). In the crystal structure of IspH from E. coli, a bound inorganic diphosphate ligand occupies the position of the diphosphate residue

Isoprenoid biosynthesis in hereditary periodic fever syndromes and inflammation

NARCIS (Netherlands)

Houten, S. M.; Frenkel, J.; Waterham, H. R.

2003-01-01

Mevalonate kinase (MK) is an essential enzyme in the isoprenoid biosynthesis pathway which produces numerous biomolecules (isoprenoids) involved in a variety of cellular processes. The indispensability of MK and isoprenoid biosynthesis for human health is demonstrated by the identification of its

The oxylipin pathway in Arabidopsis.

Science.gov (United States)

Creelman, Robert A; Mulpuri, Rao

2002-01-01

Oxylipins are acyclic or cyclic oxidation products derived from the catabolism of fatty acids which regulate many defense and developmental pathways in plants. The dramatic increase in the volume of publications and reviews on these compounds since 1997 documents the increasing interest in this compound and its role in plants. Research on this topic has solidified our understanding of the chemistry and biosynthetic pathways for oxylipin production. However, more information is still needed on how free fatty acids are produced and the role of beta-oxidation in the biosynthetic pathway for oxylipins. It is also becoming apparent that oxylipin content and composition changes during growth and development and during pathogen or insect attack. Oxylipins such as jasmonic acid (JA) or 12-oxo-phytodienoic acid modulate the expression of numerous genes and influence specific aspects of plant growth, development and responses to abiotic and biotic stresses. Although oxylipins are believed to act alone, several examples were presented to illustrate that JA-induced responses are modulated by the type and the nature of crosstalk with other signaling molecules such as ethylene and salicylic acid. How oxylipins cause changes in gene expression and instigate a physiological response is becoming understood with the isolation of mutations in both positive and negative regulators in the jasmonate signaling pathway and the use of cDNA microarrays.

Free Fatty Acids Activate Renin-Angiotensin System in 3T3-L1 Adipocytes through Nuclear Factor-kappa B Pathway

Directory of Open Access Journals (Sweden)

Jia Sun

2016-01-01

Full Text Available The activity of a local renin-angiotensin system (RAS in the adipose tissue is closely associated with obesity-related diseases. However, the mechanism of RAS activation in adipose tissue is still unknown. In the current study, we found that palmitic acid (PA, one kind of free fatty acid, induced the activity of RAS in 3T3-L1 adipocytes. In the presence of fetuin A (Fet A, PA upregulated the expression of angiotensinogen (AGT and angiotensin type 1 receptor (AT1R and stimulated the secretion of angiotensin II (ANG II in 3T3-L1 adipocytes. Moreover, the activation of RAS in 3T3-L1 adipocytes was blocked when we blocked Toll-like receptor 4 (TLR4 signaling pathway using TAK242 or NF-κB signaling pathway using BAY117082. Together, our results have identified critical molecular mechanisms linking PA/TLR4/NF-κB signaling pathway to the activity of the local renin-angiotensin system in adipose tissue.

A hepatic amino acid/mTOR/S6K-dependent signalling pathway modulates systemic lipid metabolism via neuronal signals.

Science.gov (United States)

Uno, Kenji; Yamada, Tetsuya; Ishigaki, Yasushi; Imai, Junta; Hasegawa, Yutaka; Sawada, Shojiro; Kaneko, Keizo; Ono, Hiraku; Asano, Tomoichiro; Oka, Yoshitomo; Katagiri, Hideki

2015-08-13

Metabolism is coordinated among tissues and organs via neuronal signals. Levels of circulating amino acids (AAs), which are elevated in obesity, activate the intracellular target of rapamycin complex-1 (mTORC1)/S6kinase (S6K) pathway in the liver. Here we demonstrate that hepatic AA/mTORC1/S6K signalling modulates systemic lipid metabolism via a mechanism involving neuronal inter-tissue communication. Hepatic expression of an AA transporter, SNAT2, activates the mTORC1/S6K pathway, and markedly elevates serum triglycerides (TGs), while downregulating adipose lipoprotein lipase (LPL). Hepatic Rheb or active-S6K expression have similar metabolic effects, whereas hepatic expression of dominant-negative-S6K inhibits TG elevation in SNAT2 mice. Denervation, pharmacological deafferentation and β-blocker administration suppress obesity-related hypertriglyceridemia with adipose LPL upregulation, suggesting that signals are transduced between liver and adipose tissue via a neuronal pathway consisting of afferent vagal and efferent sympathetic nerves. Thus, the neuronal mechanism uncovered here serves to coordinate amino acid and lipid levels and contributes to the development of obesity-related hypertriglyceridemia.

Reaction pathway of the degradation of the p-hydroxybenzoic acid by sulfate radical generated by ionizing radiations

International Nuclear Information System (INIS)

Criquet, Justine; Leitner, Nathalie Karpel Vel

2015-01-01

The degradation of p-hydroxybenzoic acid (HBA) in aqueous solutions by ionizing radiation was studied. The phenolic pollutant was easily removed by the electron beam irradiation, as more than 80% of the initial 100 µM introduced was degraded for a dose of 600 Gy. It was shown that the addition of persulfate, producing the sulfate radical as additional reactive species, induced a change in the reaction pathway. LC–MS analyses were performed in order to identify the different by-products formed. In the absence of persulfate, the main by-product formed was 3,4-dihydroxybenzoic acid, while in presence of persulfate, 1,4-benzoquinone was detected and the hydroxylated by-products were not present. A reaction pathway of HBA degradation by hydroxyl and sulfate radicals was proposed from the identification of the chemical structure of the different by-products detected. The influences of pH and dissolved oxygen were also studied. A high decline of HBA degradation was observed at pH 11 compared to pH 4.5, this decrease was minimized in the presence of persulfate. The dissolved oxygen concentration was found to be a limiting parameter of HBA degradation, however an excess of dissolved oxygen in solution did not improve the degradation to a large extent. - Highlights: • p-Hydroxybenzoic acid (HBA) is easily removed by e-beam irradiation. • The sulfate radicals formed from persulfate induce loss of the benzoic acid skeleton. • The dissolved oxygen concentration is a limiting parameter of the HBA degradation. • The effect of pH is minimized in presence of persulfate

Benzoic acid fermentation from starch and cellulose via a plant-like β-oxidation pathway in Streptomyces maritimus

Directory of Open Access Journals (Sweden)

Noda Shuhei

2012-04-01

Full Text Available Abstract Background Benzoic acid is one of the most useful aromatic compounds. Despite its versatility and simple structure, benzoic acid production using microbes has not been reported previously. Streptomyces are aerobic, Gram-positive, mycelia-forming soil bacteria, and are known to produce various kinds of antibiotics composed of many aromatic residues. S. maritimus possess a complex amino acid modification pathway and can serve as a new platform microbe to produce aromatic building-block compounds. In this study, we carried out benzoate fermentation using S. maritimus. In order to enhance benzoate productivity using cellulose as the carbon source, we constructed endo-glucanase secreting S. maritimus. Results After 4 days of cultivation using glucose, cellobiose, or starch as a carbon source, the maximal level of benzoate reached 257, 337, and 460 mg/l, respectively. S. maritimus expressed β-glucosidase and high amylase-retaining activity compared to those of S. lividans and S. coelicolor. In addition, for effective benzoate production from cellulosic materials, we constructed endo-glucanase-secreting S. maritimus. This transformant efficiently degraded the phosphoric acid swollen cellulose (PASC and then produced 125 mg/l benzoate. Conclusions Wild-type S. maritimus produce benzoate via a plant-like β-oxidation pathway and can assimilate various carbon sources for benzoate production. In order to encourage cellulose degradation and improve benzoate productivity from cellulose, we constructed endo-glucanase-secreting S. maritimus. Using this transformant, we also demonstrated the direct fermentation of benzoate from cellulose. To achieve further benzoate productivity, the L-phenylalanine availability needs to be improved in future.

[In vitro study over statins effects on cellular growth curves and its reversibility with mevalonate].

Science.gov (United States)

Millan Núñez-Cortés, Jesús; Alvarez Rodriguez, Ysmael; Alvarez Novés, Granada; Recarte Garcia-Andrade, Carlos; Alvarez-Sala Walther, Luis

2014-01-01

HMG-CoA-Reductase inhibitors, also known as statins, are currently the most powerful cholesterol-lowering drugs available on the market. Clinical trials and experimental evidence suggest that statins have heavy anti-atherosclerotic effects. These are in part consequence of lipid lowering but also result from pleiotropic actions of the drugs. These so-called pleiotropic properties affect various aspects of cell function, inflammation, coagulation, and vasomotor activity. These effects are mediated either indirectly through LDL-c reduction or via a direct effect on cellular functions. Although many of the pleiotropic properties of statins may be a class effect, some may be unique to certain agents and account for differences in their pharmacological activity. So, although statins typically have similar effects on LDL-c levels, differences in chemical structure and pharmacokinetic profile can lead to variations in pleiotropic effects. In this paper we analize the in vitro effects of different statins over different cell lines from cells implicated in atherosclerotic process: endothelial cells, fibroblasts, and vascular muscular cells. In relation with our results we can proof that the effects of different dosis of different statins provides singular effects over growth curves of different cellular lines, a despite of a class-dependent effects. So, pleiotropic effects and its reversibility with mevalonate are different according with the molecule and the dosis. Copyright © 2013 Elsevier España, S.L. y SEA. All rights reserved.

Nonorthologous gene displacement of phosphomevalonate kinase

NARCIS (Netherlands)

Houten, S. M.; Waterham, H. R.

2001-01-01

Phosphomevalonate kinase (PMK; EC 2.7.4.2) catalyzes the phosphorylation of 5-phosphomevalonate into 5-diphosphomevalonate, an essential step in isoprenoid biosynthesis via the mevalonate pathway. So far, two nonorthologous genes encoding PMK have been described, the Saccharomyces cerevisiae ERG8

Statin Resistance and Export

DEFF Research Database (Denmark)

Ley, Ana

Statins are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), the key enzyme in the mevalonate pathway that leads to the synthesis of cholesterol and ergosterol in animal and fungal cells, respectively. Their extensiveuse in treatment and prevention of cardiovascular diseases...

Gaseous 3-pentanol primes plant immunity against a bacterial speck pathogen, Pseudomonas syringae pv. tomato via salicylic acid and jasmonic acid-dependent signaling pathways in Arabidopsis.

Science.gov (United States)

Song, Geun C; Choi, Hye K; Ryu, Choong-Min

2015-01-01

3-Pentanol is an active organic compound produced by plants and is a component of emitted insect sex pheromones. A previous study reported that drench application of 3-pentanol elicited plant immunity against microbial pathogens and an insect pest in crop plants. Here, we evaluated whether 3-pentanol and the derivatives 1-pentanol and 2-pentanol induced plant systemic resistance using the in vitro I-plate system. Exposure of Arabidopsis seedlings to 10 μM and 100 nM 3-pentanol evaporate elicited an immune response to Pseudomonas syringae pv. tomato DC3000. We performed quantitative real-time PCR to investigate the 3-pentanol-mediated Arabidopsis immune responses by determining Pathogenesis-Related (PR) gene expression levels associated with defense signaling through salicylic acid (SA), jasmonic acid (JA), and ethylene signaling pathways. The results show that exposure to 3-pentanol and subsequent pathogen challenge upregulated PDF1.2 and PR1 expression. Selected Arabidopsis mutants confirmed that the 3-pentanol-mediated immune response involved SA and JA signaling pathways and the NPR1 gene. Taken together, this study indicates that gaseous 3-pentanol triggers induced resistance in Arabidopsis by priming SA and JA signaling pathways. To our knowledge, this is the first report that a volatile compound of an insect sex pheromone triggers plant systemic resistance against a bacterial pathogen.

Gaseous 3-pentanol primes plant immunity against a bacterial speck pathogen, Pseudomonas syringae pv. tomato via salicylic acid and jasmonic acid-dependent signaling pathways in Arabidopsis

Directory of Open Access Journals (Sweden)

Geun Cheol eSong

2015-10-01

Full Text Available 3-Pentanol is an active organic compound produced by plants and is a component of emitted insect sex pheromones. A previous study reported that drench application of 3-pentanol elicited plant immunity against microbial pathogens and an insect pest in crop plants. Here, we evaluated whether 3-pentanol and the derivatives 1-pentanol and 2-pentanol induced plant systemic resistance using the in vitro I-plate system. Exposure of Arabidopsis seedlings to 10 M and 100 nM 3-pentanol evaporate elicited an immune response to Pseudomonas syringae pv. tomato DC3000. We performed quantitative real-time PCR to investigate the 3-pentanol-mediated Arabidopsis immune responses by determining Pathogenesis-Related (PR gene expression levels associated with defense signaling through SA, JA, and ethylene signaling pathways. The results show that exposure to 3-pentanol and subsequent pathogen challenge upregulated PDF1.2 and PR1 expression. Selected Arabidopsis mutants confirmed that the 3-pentanol-mediated immune response involved salicylic acid (SA and jasmonic acid (JA signaling pathways and the NPR1 gene. Taken together, this study indicates that gaseous 3-pentanol triggers induced resistance in Arabidopsis by priming SA and JA signaling pathways. To our knowledge, this is the first report that a volatile compound of an insect sex pheromone triggers plant systemic resistance against a bacterial pathogen.

The Deoxygenation Pathways of Palmitic Acid into Hydrocarbons on Silica-Supported Ni12P5 and Ni2P Catalysts

Directory of Open Access Journals (Sweden)

Wenjun Zhou

2018-04-01

Full Text Available Pure Ni12P5/SiO2 and pure Ni2P/SiO2 catalysts were obtained by adjusting the Ni and P molar ratios, while Ni/SiO2 catalyst was prepared as a reference against which the deoxygenation pathways of palmitic acid were investigated. The catalysts were characterized by N2 adsorption, X-ray diffraction (XRD, X-ray photoelectron spectroscopy (XPS, transmission election microscopy (TEM, infrared spectroscopy of pyridine adsorption (Py-IR, H2-adsorption and temperature-programmed desorption of hydrogen (H2-TPD. The crystallographic planes of Ni(111, Ni12P5(400, Ni2P(111 were found mainly exposed on the above three catalysts, respectively. It was found that the deoxygenation pathway of palmitic acid mainly proceeded via direct decarboxylation (DCO2 to form C15 on Ni/SiO2. In contrast, on the Ni12P5/SiO2 catalyst, there were two main competitive pathways producing C15 and C16, one of which mainly proceeded via the decarbonylation (DCO to form C15 accompanying water formation, and the other pathway produced C16 via the dehydration of hexadecanol intermediate, and the yield of C15 was approximately twofold that of C16. Over the Ni2P/SiO2 catalyst, two main deoxygenation pathways formed C15, one of which was mainly the DCO pathway and the other was dehydration accompanying the hexadecanal intermediate and then direct decarbonylation without water formation. The turn over frequency (TOF followed the order: Ni12P5/SiO2 > Ni/SiO2 > Ni2P/SiO2.

Synthetic addiction extends the productive life time of engineered Escherichia coli populations

DEFF Research Database (Denmark)

Rugbjerg, Peter; Sarup-Lytzen, Kira; Nagy, Mariann

2018-01-01

range of genetic variants that disrupt the biosynthetic capacity of the engineered organism. Synthetic product addiction that couples high-yield production of a desired metabolite to expression of nonconditionally essential genes could offer a solution to this problem by selectively favoring cells...... with biosynthetic capacity in the population without constraining the medium. We constructed such synthetic product addiction by controlling the expression of two nonconditionally essential genes with a mevalonic acid biosensor. The product-addicted production organism retained high-yield mevalonic acid production...... through 95 generations of cultivation, corresponding to the number of cell generations required for >200-m3 industrial-scale production, at which time the nonaddicted strain completely abolished production. Using deep DNA sequencing, we find that the product-addicted populations do not accumulate genetic...

Amino Acids Attenuate Insulin Action on Gluconeogenesis and Promote Fatty Acid Biosynthesis via mTORC1 Signaling Pathway in trout Hepatocytes

Directory of Open Access Journals (Sweden)

Weiwei Dai

2015-06-01

Full Text Available Background/Aims: Carnivores exhibit poor utilization of dietary carbohydrates and glucose intolerant phenotypes, yet it remains unclear what are the causal factors and underlying mechanisms. We aimed to evaluate excessive amino acids (AAs-induced effects on insulin signaling, fatty acid biosynthesis and glucose metabolism in rainbow trout and determine the potential involvement of mTORC1 and p38 MAPK pathway. Methods: We stimulated trout primary hepatocytes with different AA levels and employed acute administration of rapamycin to inhibit mTORC1 activation. Results: Increased AA levels enhanced the phosphorylation of ribosomal protein S6 kinase (S6K1, S6, and insulin receptor substrate 1 (IRS-1 on Ser302 but suppressed Akt and p38 phosphorylation; up-regulated the expression of genes related to gluconeogenesis and fatty acid biosynthesis. mTORC1 inhibition not only inhibited the phosphorylation of mTORC1 downstream targets, but also blunted IRS-1 Ser302 phosphorylation and restored excessive AAs-suppressed Akt phosphorylation. Rapamycin also inhibited fatty acid biosynthetic and gluconeogenic gene expression. Conclusion: High levels of AAs up-regulate hepatic fatty acid biosynthetic gene expression through an mTORC1-dependent manner, while attenuate insulin-mediated repression of gluconeogenesis through elevating IRS-1 Ser302 phosphorylation, which in turn impairs Akt activation and thereby weakening insulin action. We propose that p38 MAPK probably also involves in these AAs-induced metabolic changes.

Toward industrial production of isoprenoids in Escherichia coli: Lessons learned from CRISPR-Cas9 based optimization of a chromosomally integrated mevalonate pathway

DEFF Research Database (Denmark)

Alonso-Gutierrez, Jorge; Koma, Daisuke; Hu, Qijun

2017-01-01

, we established a CRISPR-Cas9 system to rapidly and systematically replace promoter sequences. This strategy led to higher pathway expression and a fivefold improvement in bisabolene production. More interestingly, we analyzed proteomics data sets to understand and address some of the challenges...

Restoration of enterohepatic bile acid pathways in pregnant mice following short term activation of Fxr by GW4064

International Nuclear Information System (INIS)

Moscovitz, Jamie E.; Kong, Bo; Buckley, Kyle; Buckley, Brian; Guo, Grace L.; Aleksunes, Lauren M.

2016-01-01

The farnesoid X receptor (Fxr) controls bile acid homeostasis by coordinately regulating the expression of synthesizing enzymes (Cyp7a1, Cyp8b1), conjugating enzymes (Bal, Baat) and transporters in the ileum (Asbt, Ostα/β) and liver (Ntcp, Bsep, Ostβ). Transcriptional regulation by Fxr can be direct, or through the ileal Fgf15/FGF19 and hepatic Shp pathways. Circulating bile acids are increased during pregnancy due to hormone-mediated disruption of Fxr signaling. While this adaptation enhances lipid absorption, elevated bile acids may predispose women to develop maternal cholestasis. The objective of this study was to determine whether short-term treatment of pregnant mice with GW4064 (a potent FXR agonist) restores Fxr signaling to the level observed in virgin mice. Plasma, liver and ilea were collected from virgin and pregnant mice administered vehicle or GW4064 by oral gavage. Treatment of pregnant mice with GW4064 induced ileal Fgf15, Shp and Ostα/β mRNAs, and restored hepatic Shp, Bal, Ntcp, and Bsep back to vehicle-treated virgin levels. Pregnant mice exhibited 2.5-fold increase in Cyp7a1 mRNA compared to virgin controls, which was reduced by GW4064. Similarly treatment of mouse primary hepatocytes with plasma isolated from pregnant mice induced Cyp7a1 mRNA by nearly 3-fold as compared to virgin plasma, which could be attenuated by co-treatment with either GW4064 or recombinant FGF19 protein. Collectively, these data reveal that repressed activity of intestinal and hepatic Fxr in pregnancy, as previously demonstrated, may be restored by pharmacological activation. This study provides the basis for a novel approach to restore bile acid homeostasis in patients with maternal cholestasis. - Highlights: • Ileal bile acid pathways are altered in pregnancy in an Fxr-dependent manner. • Ileal Fxr/Fgf contributes to changes in hepatic bile acid synthesis and transport. • Treatment of pregnant mice with an Fxr agonist restores bile acid homeostasis.

Restoration of enterohepatic bile acid pathways in pregnant mice following short term activation of Fxr by GW4064

Energy Technology Data Exchange (ETDEWEB)

Moscovitz, Jamie E.; Kong, Bo; Buckley, Kyle [Department of Pharmacology and Toxicology, Rutgers University Ernest Mario School of Pharmacy, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States); Buckley, Brian [Environmental and Occupational Health Sciences Institute, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States); Guo, Grace L. [Department of Pharmacology and Toxicology, Rutgers University Ernest Mario School of Pharmacy, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States); Environmental and Occupational Health Sciences Institute, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States); Aleksunes, Lauren M., E-mail: aleksunes@eohsi.rutgers.edu [Department of Pharmacology and Toxicology, Rutgers University Ernest Mario School of Pharmacy, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States); Environmental and Occupational Health Sciences Institute, 170 Frelinghuysen Rd., Piscataway, NJ 08854 (United States)

2016-11-01

The farnesoid X receptor (Fxr) controls bile acid homeostasis by coordinately regulating the expression of synthesizing enzymes (Cyp7a1, Cyp8b1), conjugating enzymes (Bal, Baat) and transporters in the ileum (Asbt, Ostα/β) and liver (Ntcp, Bsep, Ostβ). Transcriptional regulation by Fxr can be direct, or through the ileal Fgf15/FGF19 and hepatic Shp pathways. Circulating bile acids are increased during pregnancy due to hormone-mediated disruption of Fxr signaling. While this adaptation enhances lipid absorption, elevated bile acids may predispose women to develop maternal cholestasis. The objective of this study was to determine whether short-term treatment of pregnant mice with GW4064 (a potent FXR agonist) restores Fxr signaling to the level observed in virgin mice. Plasma, liver and ilea were collected from virgin and pregnant mice administered vehicle or GW4064 by oral gavage. Treatment of pregnant mice with GW4064 induced ileal Fgf15, Shp and Ostα/β mRNAs, and restored hepatic Shp, Bal, Ntcp, and Bsep back to vehicle-treated virgin levels. Pregnant mice exhibited 2.5-fold increase in Cyp7a1 mRNA compared to virgin controls, which was reduced by GW4064. Similarly treatment of mouse primary hepatocytes with plasma isolated from pregnant mice induced Cyp7a1 mRNA by nearly 3-fold as compared to virgin plasma, which could be attenuated by co-treatment with either GW4064 or recombinant FGF19 protein. Collectively, these data reveal that repressed activity of intestinal and hepatic Fxr in pregnancy, as previously demonstrated, may be restored by pharmacological activation. This study provides the basis for a novel approach to restore bile acid homeostasis in patients with maternal cholestasis. - Highlights: • Ileal bile acid pathways are altered in pregnancy in an Fxr-dependent manner. • Ileal Fxr/Fgf contributes to changes in hepatic bile acid synthesis and transport. • Treatment of pregnant mice with an Fxr agonist restores bile acid homeostasis.

Ribosomal protein-Mdm2-p53 pathway coordinates nutrient stress with lipid metabolism by regulating MCD and promoting fatty acid oxidation.

Science.gov (United States)

Liu, Yong; He, Yizhou; Jin, Aiwen; Tikunov, Andrey P; Zhou, Lishi; Tollini, Laura A; Leslie, Patrick; Kim, Tae-Hyung; Li, Lei O; Coleman, Rosalind A; Gu, Zhennan; Chen, Yong Q; Macdonald, Jeffrey M; Graves, Lee M; Zhang, Yanping

2014-06-10

The tumor suppressor p53 has recently been shown to regulate energy metabolism through multiple mechanisms. However, the in vivo signaling pathways related to p53-mediated metabolic regulation remain largely uncharacterized. By using mice bearing a single amino acid substitution at cysteine residue 305 of mouse double minute 2 (Mdm2(C305F)), which renders Mdm2 deficient in binding ribosomal proteins (RPs) RPL11 and RPL5, we show that the RP-Mdm2-p53 signaling pathway is critical for sensing nutrient deprivation and maintaining liver lipid homeostasis. Although the Mdm2(C305F) mutation does not significantly affect growth and development in mice, this mutation promotes fat accumulation under normal feeding conditions and hepatosteatosis under acute fasting conditions. We show that nutrient deprivation inhibits rRNA biosynthesis, increases RP-Mdm2 interaction, and induces p53-mediated transactivation of malonyl-CoA decarboxylase (MCD), which catalyzes the degradation of malonyl-CoA to acetyl-CoA, thus modulating lipid partitioning. Fasted Mdm2(C305F) mice demonstrate attenuated MCD induction and enhanced malonyl-CoA accumulation in addition to decreased oxidative respiration and increased fatty acid accumulation in the liver. Thus, the RP-Mdm2-p53 pathway appears to function as an endogenous sensor responsible for stimulating fatty acid oxidation in response to nutrient depletion.

Hyaluronic acid enhances proliferation of human amniotic mesenchymal stem cells through activation of Wnt/β-catenin signaling pathway

Energy Technology Data Exchange (ETDEWEB)

Liu, Ru-Ming; Sun, Ren-Gang; Zhang, Ling-Tao; Zhang, Qing-Fang; Chen, Dai-Xiong [Guizhou Center for Translational Medicine, Affiliated Hospital of Zunyi Medical University, 149 Dalian Road, Zunyi 563000 (China); Zhong, Jian-Jiang, E-mail: jjzhong@sjtu.edu.cn [State Key Laboratory of Microbial Metabolism, and School of Life Sciences & Biotechnology, Shanghai Jiao Tong University, Shanghai 200240 (China); Xiao, Jian-Hui, E-mail: jhxiao@yahoo.com [Guizhou Center for Translational Medicine, Affiliated Hospital of Zunyi Medical University, 149 Dalian Road, Zunyi 563000 (China)

2016-07-15

This study investigated the pro-proliferative effect of hyaluronic acid (HA) on human amniotic mesenchymal stem cells (hAMSCs) and the underlying mechanisms. Treatment with HA increased cell population growth in a dose- and time-dependent manner. Analyses by flow cytometry and immunocytochemistry revealed that HA did not change the cytophenotypes of hAMSCs. Additionally, the osteogenic, chondrogenic, and adipogenic differentiation capabilities of these hAMSCs were retained after HA treatment. Moreover, HA increased the mRNA expressions of wnt1, wnt3a, wnt8a, cyclin D1, Ki-67, and β-catenin as well as the protein level of β-catenin and cyclin D1 in hAMSCs; and the nuclear localization of β-catenin was also enhanced. Furthermore, the pro-proliferative effect of HA and up-regulated expression of Wnt/β-catenin pathway-associated proteins - wnt3a, β-catenin and cyclin D1 in hAMSCs were significantly inhibited upon pre-treatment with Wnt-C59, an inhibitor of the Wnt/β-catenin pathway. These results suggest that HA may positively regulate hAMSCs proliferation through regulation of the Wnt/β-catenin signaling pathway. - Highlights: • Hyaluronic acid (HA) could promote the proliferation of hAMSCs. • HA treatment dose not affect the pluripotency of hAMSCs. • HA increases hAMSCs proliferation through activation of Wnt/β-catenin signaling.

Cross-talk between abscisic acid-dependent and abscisic acid-independent pathways during abiotic stress.

Science.gov (United States)

Roychoudhury, Aryadeep; Paul, Saikat; Basu, Supratim

2013-07-01

Salinity, drought and low temperature are the common forms of abiotic stress encountered by land plants. To cope with these adverse environmental factors, plants execute several physiological and metabolic responses. Both osmotic stress (elicited by water deficit or high salt) and cold stress increase the endogenous level of the phytohormone abscisic acid (ABA). ABA-dependent stomatal closure to reduce water loss is associated with small signaling molecules like nitric oxide, reactive oxygen species and cytosolic free calcium, and mediated by rapidly altering ion fluxes in guard cells. ABA also triggers the expression of osmotic stress-responsive (OR) genes, which usually contain single/multiple copies of cis-acting sequence called abscisic acid-responsive element (ABRE) in their upstream regions, mostly recognized by the basic leucine zipper-transcription factors (TFs), namely, ABA-responsive element-binding protein/ABA-binding factor. Another conserved sequence called the dehydration-responsive element (DRE)/C-repeat, responding to cold or osmotic stress, but not to ABA, occurs in some OR promoters, to which the DRE-binding protein/C-repeat-binding factor binds. In contrast, there are genes or TFs containing both DRE/CRT and ABRE, which can integrate input stimuli from salinity, drought, cold and ABA signaling pathways, thereby enabling cross-tolerance to multiple stresses. A strong candidate that mediates such cross-talk is calcium, which serves as a common second messenger for abiotic stress conditions and ABA. The present review highlights the involvement of both ABA-dependent and ABA-independent signaling components and their interaction or convergence in activating the stress genes. We restrict our discussion to salinity, drought and cold stress.

Retinoic acid activates two pathways required for meiosis in mice.

Directory of Open Access Journals (Sweden)

Jana Koubova

2014-08-01

Full Text Available In all sexually reproducing organisms, cells of the germ line must transition from mitosis to meiosis. In mice, retinoic acid (RA, the extrinsic signal for meiotic initiation, activates transcription of Stra8, which is required for meiotic DNA replication and the subsequent processes of meiotic prophase. Here we report that RA also activates transcription of Rec8, which encodes a component of the cohesin complex that accumulates during meiotic S phase, and which is essential for chromosome synapsis and segregation. This RA induction of Rec8 occurs in parallel with the induction of Stra8, and independently of Stra8 function, and it is conserved between the sexes. Further, RA induction of Rec8, like that of Stra8, requires the germ-cell-intrinsic competence factor Dazl. Our findings strengthen the importance of RA and Dazl in the meiotic transition, provide important details about the Stra8 pathway, and open avenues to investigate early meiosis through analysis of Rec8 induction and function.

Differential effects of 17 alpha-ethinylestradiol on the neutral and acidic pathways of bile salt synthesis in the rat

NARCIS (Netherlands)

Koopen, NR; Post, SM; Wolters, H; Havinga, R; Stellaard, F; Boverhof, R; Kuipers, F; Princen, HMG

Effects of 17 alpha-ethinylestradiol (EE) on the neutral and acidic biosynthetic pathways of bile salt (BS) synthesis were evaluated in rats with an intact enterohepatic circulation and in rats with long-term bile diversion to induce BS synthesis, For this purpose, bile salt pool composition,

All-trans retinoic acid promotes neural lineage entry by pluripotent embryonic stem cells via multiple pathways

Directory of Open Access Journals (Sweden)

Fang Bo

2009-07-01

Full Text Available Abstract Background All-trans retinoic acid (RA is one of the most important morphogens with pleiotropic actions. Its embryonic distribution correlates with neural differentiation in the developing central nervous system. To explore the precise effects of RA on neural differentiation of mouse embryonic stem cells (ESCs, we detected expression of RA nuclear receptors and RA-metabolizing enzymes in mouse ESCs and investigated the roles of RA in adherent monolayer culture. Results Upon addition of RA, cell differentiation was directed rapidly and exclusively into the neural lineage. Conversely, pharmacological interference with RA signaling suppressed this neural differentiation. Inhibition of fibroblast growth factor (FGF signaling did not suppress significantly neural differentiation in RA-treated cultures. Pharmacological interference with extracellular signal-regulated kinase (ERK pathway or activation of Wnt pathway effectively blocked the RA-promoted neural specification. ERK phosphorylation was enhanced in RA-treated cultures at the early stage of differentiation. Conclusion RA can promote neural lineage entry by ESCs in adherent monolayer culture systems. This effect depends on RA signaling and its crosstalk with the ERK and Wnt pathways.

Detection of phytohormones in temperate forest fungi predicts consistent abscisic acid production and a common pathway for cytokinin biosynthesis.

Science.gov (United States)

Morrison, Erin N; Knowles, Sarah; Hayward, Allison; Thorn, R Greg; Saville, Barry J; Emery, R J N

2015-01-01

The phytohormones, abscisic acid and cytokinin, once were thought to be present uniquely in plants, but increasing evidence suggests that these hormones are present in a wide variety of organisms. Few studies have examined fungi for the presence of these "plant" hormones or addressed whether their levels differ based on the nutrition mode of the fungus. This study examined 20 temperate forest fungi of differing nutritional modes (ectomycorrhizal, wood-rotting, saprotrophic). Abscisic acid and cytokinin were present in all fungi sampled; this indicated that the sampled fungi have the capacity to synthesize these two classes of phytohormones. Of the 27 cytokinins analyzed by HPLC-ESI MS/MS, seven were present in all fungi sampled. This suggested the existence of a common cytokinin metabolic pathway in fungi that does not vary among different nutritional modes. Predictions regarding the source of isopentenyl, cis-zeatin and methylthiol CK production stemming from the tRNA degradation pathway among fungi are discussed. © 2015 by The Mycological Society of America.

Metabolic pathway engineering based on metabolomics confers acetic and formic acid tolerance to a recombinant xylose-fermenting strain of Saccharomyces cerevisiae

Directory of Open Access Journals (Sweden)

Ishii Jun

2011-01-01

Full Text Available Abstract Background The development of novel yeast strains with increased tolerance toward inhibitors in lignocellulosic hydrolysates is highly desirable for the production of bio-ethanol. Weak organic acids such as acetic and formic acids are necessarily released during the pretreatment (i.e. solubilization and hydrolysis of lignocelluloses, which negatively affect microbial growth and ethanol production. However, since the mode of toxicity is complicated, genetic engineering strategies addressing yeast tolerance to weak organic acids have been rare. Thus, enhanced basic research is expected to identify target genes for improved weak acid tolerance. Results In this study, the effect of acetic acid on xylose fermentation was analyzed by examining metabolite profiles in a recombinant xylose-fermenting strain of Saccharomyces cerevisiae. Metabolome analysis revealed that metabolites involved in the non-oxidative pentose phosphate pathway (PPP [e.g. sedoheptulose-7-phosphate, ribulose-5-phosphate, ribose-5-phosphate and erythrose-4-phosphate] were significantly accumulated by the addition of acetate, indicating the possibility that acetic acid slows down the flux of the pathway. Accordingly, a gene encoding a PPP-related enzyme, transaldolase or transketolase, was overexpressed in the xylose-fermenting yeast, which successfully conferred increased ethanol productivity in the presence of acetic and formic acid. Conclusions Our metabolomic approach revealed one of the molecular events underlying the response to acetic acid and focuses attention on the non-oxidative PPP as a target for metabolic engineering. An important challenge for metabolic engineering is identification of gene targets that have material importance. This study has demonstrated that metabolomics is a powerful tool to develop rational strategies to confer tolerance to stress through genetic engineering.

The effect of MEP pathway and other inhibitors on the intracellular localization of a plasma membrane-targeted, isoprenylable GFP reporter protein in tobacco BY-2 cells [v1; ref status: indexed, http://f1000r.es/yx

Directory of Open Access Journals (Sweden)

Michael Hartmann

2013-08-01

Full Text Available We have established an in vivo visualization system for the geranylgeranylation of proteins in a stably transformed tobacco BY-2 cell line, based on the expression of a dexamethasone-inducible GFP fused to the carboxy-terminal basic domain of the rice calmodulin CaM61, which naturally bears a CaaL geranylgeranylation motif (GFP-BD-CVIL. By using pathway-specific inhibitors it was demonstrated that inhibition of the methylerythritol phosphate (MEP pathway with known inhibitors like oxoclomazone and fosmidomycin, as well as inhibition of the protein geranylgeranyltransferase type 1 (PGGT-1, shifted the localization of the GFP-BD-CVIL protein from the membrane to the nucleus. In contrast, the inhibition of the mevalonate (MVA pathway with mevinolin did not affect the localization. During the present work, this test system has been used to examine the effect of newly designed inhibitors of the MEP pathway and inhibitors of sterol biosynthesis such as squalestatin, terbinafine and Ro48-8071. In addition, we also studied the impact of different post-prenylation inhibitors or those suspected to affect the transport of proteins to the plasma membrane on the localization of the geranylgeranylable fusion protein GFP-BD-CVIL.

The intracellular cholesterol landscape: dynamic integrator of the immune response

Science.gov (United States)

Fessler, Michael B.

2016-01-01

Cholesterol has typically been considered an exogenous, disease-related factor in immunity; however, recent literature suggests that a paradigm shift is in order. Sterols are now recognized to ligate several immune receptors. Altered flux through the mevalonic acid synthesis pathway also appears to be a required event in the antiviral interferon response of macrophages and in the activation, proliferation, and differentiation of T cells. In this review, evidence is discussed that suggests an intrinsic, ‘professional’ role for sterols and oxysterols in macrophage and T cell immunity. Host defense may have been the original selection pressure behind the development of mechanisms for intracellular cholesterol homeostasis. Functional coupling between sterol metabolism and immunity has fundamental implications for health and disease. PMID:27692616

Targeting the Mevalonate Pathway to Reduce Mortality from Ovarian Cancer

Science.gov (United States)

2015-10-01

RHOB, RRAS, RHOA, PLAU, RHOF, ITGB5, ITGB3 Methylglyoxal degradation III 2.17Eþ00 1.30E01 AKR1C1/AKR1C2, AKR1C3, AKR1C4 Dopamine degradation...2007;25: 2921–7. 6. Bober SL, Recklitis CJ, Bakan J, Garber JE, Patenaude AF. Addressing sexual dysfunction after risk-reducing salpingo-oophorectomy

Targeting the Mevalonate Pathway to Reduce Mortality from Ovarian Cancer

Science.gov (United States)

2017-12-01

removed by Picard Tools, and base quality score recalibration and Indel (insert/deletion) realignment using the Genome Analysis Toolkit GATK (McKenna et...Analysis Toolkit : a MapReduce framework for analyzing next-generation DNA sequencing data. Genome research 20, 1297-1303. McLean, C. Y., Bristor, D...CA151683), a Department of Defense Career Development award (CA130247), and grants from Gabrielle’s Angel Foundation and Concern Foundation. GGW is also

Targeting the Mevalonate Pathway to Reduce Mortality from Ovarian Cancer

Science.gov (United States)

2016-10-01

Register Center (information on born children and 1st level female siblings , emigration and death); and 4) patient register, HILMO (information on co...dose and duration, timing of the intervention (pre vs . post diagnosis use), histologic subtype, and patterns of adherence after adjusting for...duration, timing of the intervention (pre vs . post diagnosis use), histologic subtype, and patterns of adherence after adjusting for prespecified

De novo assembly and functional annotation of Myrciaria dubia fruit transcriptome reveals multiple metabolic pathways for L-ascorbic acid biosynthesis.

Science.gov (United States)

Castro, Juan C; Maddox, J Dylan; Cobos, Marianela; Requena, David; Zimic, Mirko; Bombarely, Aureliano; Imán, Sixto A; Cerdeira, Luis A; Medina, Andersson E

2015-11-24

Myrciaria dubia is an Amazonian fruit shrub that produces numerous bioactive phytochemicals, but is best known by its high L-ascorbic acid (AsA) content in fruits. Pronounced variation in AsA content has been observed both within and among individuals, but the genetic factors responsible for this variation are largely unknown. The goals of this research, therefore, were to assemble, characterize, and annotate the fruit transcriptome of M. dubia in order to reconstruct metabolic pathways and determine if multiple pathways contribute to AsA biosynthesis. In total 24,551,882 high-quality sequence reads were de novo assembled into 70,048 unigenes (mean length = 1150 bp, N50 = 1775 bp). Assembled sequences were annotated using BLASTX against public databases such as TAIR, GR-protein, FB, MGI, RGD, ZFIN, SGN, WB, TIGR_CMR, and JCVI-CMR with 75.2 % of unigenes having annotations. Of the three core GO annotation categories, biological processes comprised 53.6 % of the total assigned annotations, whereas cellular components and molecular functions comprised 23.3 and 23.1 %, respectively. Based on the KEGG pathway assignment of the functionally annotated transcripts, five metabolic pathways for AsA biosynthesis were identified: animal-like pathway, myo-inositol pathway, L-gulose pathway, D-mannose/L-galactose pathway, and uronic acid pathway. All transcripts coding enzymes involved in the ascorbate-glutathione cycle were also identified. Finally, we used the assembly to identified 6314 genic microsatellites and 23,481 high quality SNPs. This study describes the first next-generation sequencing effort and transcriptome annotation of a non-model Amazonian plant that is relevant for AsA production and other bioactive phytochemicals. Genes encoding key enzymes were successfully identified and metabolic pathways involved in biosynthesis of AsA, anthocyanins, and other metabolic pathways have been reconstructed. The identification of these genes and pathways is in agreement with

Drosophila fatty acid taste signals through the PLC pathway in sugar-sensing neurons.

Directory of Open Access Journals (Sweden)

Pavel Masek

Full Text Available Taste is the primary sensory system for detecting food quality and palatability. Drosophila detects five distinct taste modalities that include sweet, bitter, salt, water, and the taste of carbonation. Of these, sweet-sensing neurons appear to have utility for the detection of nutritionally rich food while bitter-sensing neurons signal toxicity and confer repulsion. Growing evidence in mammals suggests that taste for fatty acids (FAs signals the presence of dietary lipids and promotes feeding. While flies appear to be attracted to fatty acids, the neural basis for fatty acid detection and attraction are unclear. Here, we demonstrate that a range of FAs are detected by the fly gustatory system and elicit a robust feeding response. Flies lacking olfactory organs respond robustly to FAs, confirming that FA attraction is mediated through the gustatory system. Furthermore, flies detect FAs independent of pH, suggesting the molecular basis for FA taste is not due to acidity. We show that low and medium concentrations of FAs serve as an appetitive signal and they are detected exclusively through the same subset of neurons that sense appetitive sweet substances, including most sugars. In mammals, taste perception of sweet and bitter substances is dependent on phospholipase C (PLC signaling in specialized taste buds. We find that flies mutant for norpA, a Drosophila ortholog of PLC, fail to respond to FAs. Intriguingly, norpA mutants respond normally to other tastants, including sucrose and yeast. The defect of norpA mutants can be rescued by selectively restoring norpA expression in sweet-sensing neurons, corroborating that FAs signal through sweet-sensing neurons, and suggesting PLC signaling in the gustatory system is specifically involved in FA taste. Taken together, these findings reveal that PLC function in Drosophila sweet-sensing neurons is a conserved molecular signaling pathway that confers attraction to fatty acids.

Relevance of DNA repair pathways on ascorbic acid effects on Echerichia Coli K-12 cells

International Nuclear Information System (INIS)

Slyus, M.A. van; Oliveira, R.L.B. da C.; Felzenszwalb, I.; Gomes, R.A.; Menck, C.F.

1985-01-01

Inactivation kinetics were performed with repair proficient and deficient Escherichia coli K-12 cells treated with oxidized solutions of ascorbic acid. The repair pathways controlled by the recA and uvrA gene products are essential for cell survival to the treatment. However, SOS chromotest result indicates that the SOS functions are only induced at high and toxic concentrations of the drug. Moreover, single strand breaks in DNA from treated cells are detected, demonstrating genome damage promoted by oxidized solutions of ascorbate. (M.A.C.) [pt

On the mechanism for the formation of indole alkalois in Penicillium concavo-rugulosum

International Nuclear Information System (INIS)

Ohmomo, Sadahiro; Miyazaki, Kenji; Ohashi, Tsutomu; Abe, Matazo

1977-01-01

Experiments on the biosynthesis and microbiological conversion of indole alkaloids in Penicillium concavo-rugulosum were carried out with the growing and resting mycelia, respectively, of a selected strain of the same mold. The former experiments were performed by the use of DL-tryptophan-3- 14 C or DL-mevalonic acid-2- 14 C-lactone as a precursor, while the latter experiments by the use of rugulovasine A- 3 H, dihydrorugulovasine A- 3 H, 4-[γ,γ-dimethylallyl]-tryptophan- 3 H, chanoclavine-[I]- 3 H or the other tritiated ergoline alkaloids. The results of these experiments suggested that in the Penicillium mold employed there exist the following biosynthetic route: tryptophan+mevalonic acid → 4-[γ,γ-dimethylallyl]-tryptophan → rugulovasine A → dihydrorugulovasine A → dihydrorugulovasine A-lactam. (auth.)

Heterologous expression of the isopimaric acid pathway in Nicotiana benthamiana and the effect of N-terminal modifications of the involved cytochrome P450 enzyme

DEFF Research Database (Denmark)

Gnanasekaran, Thiyagarajan; Vavitsas, Konstantinos; Andersen-Ranberg, Johan

2015-01-01

in the infiltrated leaves. Furthermore, we demonstrated that a modified membrane anchor is a prerequisite for a functional CYP720B4 enzyme when the chloroplast targeting peptide is added. We report the accumulation of 45-55 μg/g plant dry weight of isopimaric acid four days after the infiltration with the modified...... in the chloroplast and subsequently oxidized by a cytochrome P450, CYP720B4. RESULTS: We transiently expressed the isopimaric acid pathway in Nicotiana benthamiana leaves and enhanced its productivity by the expression of two rate-limiting steps in the pathway (providing the general precursor of diterpenes). This co...

Evidence of a New Role for the High-Osmolarity Glycerol Mitogen-Activated Protein Kinase Pathway in Yeast: Regulating Adaptation to Citric Acid Stress†

OpenAIRE

Lawrence, Clare L.; Botting, Catherine H.; Antrobus, Robin; Coote, Peter J.

2004-01-01

Screening the Saccharomyces cerevisiae disruptome, profiling transcripts, and determining changes in protein expression have identified an important new role for the high-osmolarity glycerol (HOG) mitogen-activated protein kinase (MAPK) pathway in the regulation of adaptation to citric acid stress. Deletion of HOG1, SSK1, PBS2, PTC2, PTP2, and PTP3 resulted in sensitivity to citric acid. Furthermore, citric acid resulted in the dual phosphorylation, and thus activation, of Hog1p. Despite mino...

Effects of EPSPS Copy Number Variation (CNV and Glyphosate Application on the Aromatic and Branched Chain Amino Acid Synthesis Pathways in Amaranthus palmeri

Directory of Open Access Journals (Sweden)

Manuel Fernández-Escalada

2017-11-01

Full Text Available A key enzyme of the shikimate pathway, 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS; EC 2.5.1.19, is the known target of the widely used herbicide glyphosate. Glyphosate resistance in Amaranthus palmeri, one of the most troublesome weeds in agriculture, has evolved through increased EPSPS gene copy number. The aim of this work was to study the pleiotropic effects of (i EPSPS increased transcript abundance due to gene copy number variation (CNV and of (ii glyphosate application on the aromatic amino acid (AAA and branched chain amino acid (BCAA synthesis pathways. Hydroponically grown glyphosate sensitive (GS and glyphosate resistant (GR plants were treated with glyphosate 3 days after treatment. In absence of glyphosate treatment, high EPSPS gene copy number had only a subtle effect on transcriptional regulation of AAA and BCAA pathway genes. In contrast, glyphosate treatment provoked a general accumulation of the transcripts corresponding to genes of the AAA pathway leading to synthesis of chorismate in both GS and GR. After chorismate, anthranilate synthase transcript abundance was higher while chorismate mutase transcription showed a small decrease in GR and remained stable in GS, suggesting a regulatory branch point in the pathway that favors synthesis toward tryptophan over phenylalanine and tyrosine after glyphosate treatment. This was confirmed by studying enzyme activities in vitro and amino acid analysis. Importantly, this upregulation was glyphosate dose dependent and was observed similarly in both GS and GR populations. Glyphosate treatment also had a slight effect on the expression of BCAA genes but no general effect on the pathway could be observed. Taken together, our observations suggest that the high CNV of EPSPS in A. palmeri GR populations has no major pleiotropic effect on the expression of AAA biosynthetic genes, even in response to glyphosate treatment. This finding supports the idea that the fitness cost associated

Covalent bonding of chloroanilines to humic constituents: Pathways, kinetics, and stability

International Nuclear Information System (INIS)

Kong, Deyang; Xia, Qing; Liu, Guoqiang; Huang, Qingguo; Lu, Junhe

2013-01-01

Covalent coupling to natural humic constituents comprises an important transformation pathway for anilinic pollutants in the environment. We systematically investigated the reactions of chlorine substituted anilines with catechol and syringic acid in horseradish peroxidase (HRP) catalyzed systems. It was demonstrated that although nucleophilic addition was the mechanism of covalent bonding to both catechol and syringic acid, chloroanilines coupled to the 2 humic constituents via slightly different pathways. 1,4-addition and 1,2-addition are involved to catechol and syringic acid, respectively. 1,4-addition showed empirical 2nd order kinetics and this pathway seemed to be more permanent than 1,2-addition. Stability experiments demonstrated that cross-coupling products with syringic acid could be easily released in acidic conditions. However, cross-coupling with catechol was relatively stable at similar conditions. Thus, the environmental behavior and bioavailability of the coupling products should be carefully assessed. -- Highlights: •Chloroanilines covalently coupled to humic constituents in HRP catalyzed processes, which facilitated their transformation. •MS technique was employed to analyze the coupling products and therefore elucidate the reaction pathways. •Chloroanilines couple to catechol and syringic acid via 1,4- and 1,2-nucleophilic addition pathways, respectively. •Cross-coupling products formed via 1,4-nucleophilic addition pathway were more stable than those via 1,2-addition pathway. -- Bound residues of chloroanilines formed via 1,2- and 1,4-nucleophilic addition pathways showed different stability

Inhibition of Melanogenesis by Gallic Acid: Possible Involvement of the PI3K/Akt, MEK/ERK and Wnt/β-Catenin Signaling Pathways in B16F10 Cells

Science.gov (United States)

Su, Tzu-Rong; Lin, Jen-Jie; Tsai, Chi-Chu; Huang, Tsu-Kei; Yang, Zih-Yan; Wu, Ming-O; Zheng, Yu-Qing; Su, Ching-Chyuan; Wu, Yu-Jen

2013-01-01

Gallic acid is one of the major flavonoids found in plants. It acts as an antioxidant, and seems to have anti-inflammatory, anti-viral, and anti-cancer properties. In this study, we investigated the effects of gallic acid on melanogenesis, including the activation of melanogenesis signaling pathways. Gallic acid significantly inhibited both melanin synthesis and tyrosinase activity in a dose- and time-dependent manner, and decreased the expression of melanogenesis-related proteins, such as microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP1), and dopachrome tautomerase (Dct). In addition, gallic acid also acts by phosphorylating and activating melanogenesis inhibitory proteins such as Akt and mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK). Using inhibitors against PI3K/Akt (LY294002) or MEK/ERK-specific (PD98059), the hypopigmentation effect was suppressed, and the gallic acid-initiated activation of MEK/ERK and PI3K/Akt was also revoked. Gallic acid also increased GSK3β and p-β-catenin expression but down-regulated p-GSK3β. Moreover, GSK3β-specific inhibitor (SB216763) restored gallic acid-induced melanin reduction. These results suggest that activation of the MEK/ERK, PI3K/Akt, and inhibition of Wnt/β-catenin signaling pathways is involved in the melanogenesis signaling cascade, and that activation by gallic acid reduces melanin synthesis via down-regulation of MITF and its downstream signaling pathway. In conclusion, gallic acid may be a potentially agent for the treatment of certain skin conditions. PMID:24129178

Inhibition of Melanogenesis by Gallic Acid: Possible Involvement of the PI3K/Akt, MEK/ERK and Wnt/β-Catenin Signaling Pathways in B16F10 Cells

Directory of Open Access Journals (Sweden)

Yu-Jen Wu

2013-10-01

Full Text Available Gallic acid is one of the major flavonoids found in plants. It acts as an antioxidant, and seems to have anti-inflammatory, anti-viral, and anti-cancer properties. In this study, we investigated the effects of gallic acid on melanogenesis, including the activation of melanogenesis signaling pathways. Gallic acid significantly inhibited both melanin synthesis and tyrosinase activity in a dose- and time-dependent manner, and decreased the expression of melanogenesis-related proteins, such as microphthalmia-associated transcription factor (MITF, tyrosinase, tyrosinase-related protein-1 (TRP1, and dopachrome tautomerase (Dct. In addition, gallic acid also acts by phosphorylating and activating melanogenesis inhibitory proteins such as Akt and mitogen-activated protein kinase (MEK/extracellular signal-regulated kinase (ERK. Using inhibitors against PI3K/Akt (LY294002 or MEK/ERK-specific (PD98059, the hypopigmentation effect was suppressed, and the gallic acid-initiated activation of MEK/ERK and PI3K/Akt was also revoked. Gallic acid also increased GSK3β and p-β-catenin expression but down-regulated p-GSK3β. Moreover, GSK3β-specific inhibitor (SB216763 restored gallic acid-induced melanin reduction. These results suggest that activation of the MEK/ERK, PI3K/Akt, and inhibition of Wnt/β-catenin signaling pathways is involved in the melanogenesis signaling cascade, and that activation by gallic acid reduces melanin synthesis via down-regulation of MITF and its downstream signaling pathway. In conclusion, gallic acid may be a potentially agent for the treatment of certain skin conditions.

Modulation of Immunological Pathways in Autistic and Neurotypical Lymphoblastoid Cell Lines by the Enteric Microbiome Metabolite Propionic Acid.

Science.gov (United States)

Frye, Richard E; Nankova, Bistra; Bhattacharyya, Sudeepa; Rose, Shannon; Bennuri, Sirish C; MacFabe, Derrick F

2017-01-01

Propionic acid (PPA) is a ubiquitous short-chain fatty acid which is a fermentation product of the enteric microbiome and present or added to many foods. While PPA has beneficial effects, it is also associated with human disorders, including autism spectrum disorders (ASDs). We previously demonstrated that PPA modulates mitochondrial dysfunction differentially in subsets of lymphoblastoid cell lines (LCLs) derived from patients with ASD. Specifically, PPA significantly increases mitochondrial function in LCLs that have mitochondrial dysfunction at baseline [individuals with autistic disorder with atypical mitochondrial function (AD-A) LCLs] as compared to ASD LCLs with normal mitochondrial function [individuals with autistic disorder with normal mitochondrial function (AD-N) LCLs] and control (CNT) LCLs. PPA at 1 mM was found to have a minimal effect on expression of immune genes in CNT and AD-N LCLs. However, as hypothesized, Panther analysis demonstrated that 1 mM PPA exposure at 24 or 48 h resulted in significant activation of the immune system genes in AD-A LCLs. When the effect of PPA on ASD LCLs were compared to the CNT LCLs, both ASD groups demonstrated immune pathway activation, although the AD-A LCLs demonstrate a wider activation of immune genes. Ingenuity Pathway Analysis identified several immune-related pathways as key Canonical Pathways that were differentially regulated, specifically human leukocyte antigen expression and immunoglobulin production genes were upregulated. These data demonstrate that the enteric microbiome metabolite PPA can evoke atypical immune activation in LCLs with an underlying abnormal metabolic state. As PPA, as well as enteric bacteria which produce PPA, have been implicated in a wide variety of diseases which have components of immune dysfunction, including ASD, diabetes, obesity, and inflammatory diseases, insight into this metabolic modulator may have wide applications for both health and disease.

Proteomic Analysis of Lettuce Seed Germination and Thermoinhibition by Sampling of Individual Seeds at Germination and Removal of Storage Proteins by Polyethylene Glycol Fractionation1

Science.gov (United States)

Song, Bin-Yan; Deng, Zhi-Jun; Wang, Yue; Liu, Shu-Jun; Møller, Ian Max; Song, Song-Quan

2015-01-01

Germination and thermoinhibition in lettuce (Lactuca sativa ‘Jianyexianfeng No. 1’) seeds were investigated by a proteomic comparison among dry seeds, germinated seeds at 15°C, at 15°C after imbibition at 25°C for 48 h, or at 25°C in KNO3 (all sampled individually at germination), and ungerminated seeds at 25°C, a thermoinhibitory temperature. Before two-dimensional gel electrophoresis analysis, storage proteins (greater than 50% of total extractable protein) were removed by polyethylene glycol precipitation, which significantly improved the detection of less abundant proteins on two-dimensional gels. A total of 108 protein spots were identified to change more than 2-fold (P lettuce seed germination and thermoinhibition. Accumulation of three proteins and expression of five genes participating in the mevalonate (MVA) pathway of isoprenoid biosynthesis correlated positively with seed germinability. Inhibition of this pathway by lovastatin delayed seed germination and increased the sensitivity of germination to abscisic acid. MVA pathway-derived products, cytokinins, partially reversed the lovastatin inhibition of germination and released seed thermoinhibition at 25°C. We conclude that the MVA pathway for isoprenoid biosynthesis is involved in lettuce seed germination and thermoinhibition. PMID:25736209

Biosynthesis of furanochromones in Pimpinella monoica

Indian Academy of Sciences (India)

polyketide origin of their aromatic and pyrone rings while the furan ring originates via an acetate-mevalonate pathway. The plant also utilises glycine and leucine as substrate via acetate. Biotransformation of 3-H-visnagin to (6) but not to (2) was also observed. Keywords. Biosynthesis; furochromones; polyketide origin; ...

Amino acid regulation of autophagosome formation

NARCIS (Netherlands)

Meijer, Alfred J.

2008-01-01

Amino acids are not only substrates for various metabolic pathways, but can also serve as signaling molecules controlling signal transduction pathways. One of these signaling pathways is mTOR-dependent and is activated by amino acids (leucine in particular) in synergy with insulin. Activation of

Uric acid stimulates proliferative pathways in vascular smooth muscle cells through the activation of p38 MAPK, p44/42 MAPK and PDGFRβ.

Science.gov (United States)

Kırça, M; Oğuz, N; Çetin, A; Uzuner, F; Yeşilkaya, A

2017-04-01

Hyperuricemia and angiotensin II (Ang II) may have a pathogenetic role in the development of hypertension and atherosclerosis as well as cardiovascular disease (CVD) and its prognosis. The purpose of this study was to investigate whether uric acid can induce proliferative pathways of vascular smooth muscle cell (VSMC) that are thought to be responsible for the development of CVD. The phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), p44/42 mitogen-activated protein kinase (p44/42 MAPK) and platelet-derived growth factor receptor β (PDGFRβ) was measured by Elisa and Western blot techniques to determine the activation of proliferative pathways in primary cultured VSMCs from rat aorta. Results demonstrated that uric acid can stimulate p38 MAPK, p44/42 MAPK and PDGFRβ phosphorylation in a time- and concentration-dependent manner. Furthermore, treatment of VSMCs with the angiotensin II type I receptor (AT1R) inhibitor losartan suppressed p38 MAPK and p44/42 MAPK induction by uric acid. The stimulatory effect of uric acid on p38 MAPK was higher compared to that of Ang II. The results of this study show for the first time that uric acid-induced PDGFRβ phosphorylation plays a crucial role in the development of CVDs and that elevated uric acid levels could be a potential therapeutical target in CVD patients.

Differential regulation of EGFR-MAPK signaling by deoxycholic acid (DCA) and ursodeoxycholic acid (UDCA) in colon cancer.

Science.gov (United States)

Centuori, Sara M; Martinez, Jesse D

2014-10-01

A high-fat diet coincides with increased levels of bile acids. This increase in bile acids, particularly deoxycholic acid (DCA), has been strongly associated with the development of colon cancer. Conversely, ursodeoxycholic acid (UDCA) may have chemopreventive properties. Although structurally similar, DCA and UDCA present different biological and pathological effects in colon cancer progression. The differential regulation of cancer by these two bile acids is not yet fully understood. However, one possible explanation for their diverging effects is their ability to differentially regulate signaling pathways involved in the multistep progression of colon cancer, such as the epidermal growth factor receptor (EGFR)-mitogen-activated protein kinase (MAPK) pathway. This review will examine the biological effects of DCA and UDCA on colon cancer development, as well as the diverging effects of these bile acids on the oncogenic signaling pathways that play a role in colon cancer development, with a particular emphasis on bile acid regulation of the EGFR-MAPK pathway.

The Plastidial 2-C-Methyl-d-Erythritol 4-Phosphate Pathway Provides the Isoprenyl Moiety for Protein Geranylgeranylation in Tobacco BY-2 Cells[C][W

Science.gov (United States)

Gerber, Esther; Hemmerlin, Andréa; Hartmann, Michael; Heintz, Dimitri; Hartmann, Marie-Andrée; Mutterer, Jérôme; Rodríguez-Concepción, Manuel; Boronat, Albert; Van Dorsselaer, Alain; Rohmer, Michel; Crowell, Dring N.; Bach, Thomas J.

2009-01-01

Protein farnesylation and geranylgeranylation are important posttranslational modifications in eukaryotic cells. We visualized in transformed Nicotiana tabacum Bright Yellow-2 (BY-2) cells the geranylgeranylation and plasma membrane localization of GFP-BD-CVIL, which consists of green fluorescent protein (GFP) fused to the C-terminal polybasic domain (BD) and CVIL isoprenylation motif from the Oryza sativa calmodulin, CaM61. Treatment with fosmidomycin (Fos) or oxoclomazone (OC), inhibitors of the plastidial 2-C-methyl-d-erythritol 4-phosphate (MEP) pathway, caused mislocalization of the protein to the nucleus, whereas treatment with mevinolin, an inhibitor of the cytosolic mevalonate pathway, did not. The nuclear localization of GFP-BD-CVIL in the presence of MEP pathway inhibitors was completely reversed by all-trans-geranylgeraniol (GGol). Furthermore, 1-deoxy-d-xylulose (DX) reversed the effects of OC, but not Fos, consistent with the hypothesis that OC blocks 1-deoxy-d-xylulose 5-phosphate synthesis, whereas Fos inhibits its conversion to 2-C-methyl-d-erythritol 4-phosphate. By contrast, GGol and DX did not rescue the nuclear mislocalization of GFP-BD-CVIL in the presence of a protein geranylgeranyltransferase type 1 inhibitor. Thus, the MEP pathway has an essential role in geranylgeranyl diphosphate (GGPP) biosynthesis and protein geranylgeranylation in BY-2 cells. GFP-BD-CVIL is a versatile tool for identifying pharmaceuticals and herbicides that interfere either with GGPP biosynthesis or with protein geranylgeranylation. PMID:19136647

Microbial Propionic Acid Production

Directory of Open Access Journals (Sweden)

R. Axayacatl Gonzalez-Garcia

2017-05-01

Full Text Available Propionic acid (propionate is a commercially valuable carboxylic acid produced through microbial fermentation. Propionic acid is mainly used in the food industry but has recently found applications in the cosmetic, plastics and pharmaceutical industries. Propionate can be produced via various metabolic pathways, which can be classified into three major groups: fermentative pathways, biosynthetic pathways, and amino acid catabolic pathways. The current review provides an in-depth description of the major metabolic routes for propionate production from an energy optimization perspective. Biological propionate production is limited by high downstream purification costs which can be addressed if the target yield, productivity and titre can be achieved. Genome shuffling combined with high throughput omics and metabolic engineering is providing new opportunities, and biological propionate production is likely to enter the market in the not so distant future. In order to realise the full potential of metabolic engineering and heterologous expression, however, a greater understanding of metabolic capabilities of the native producers, the fittest producers, is required.

Enhanced accumulation of phytosterols and phenolic compounds in cyclodextrin-elicited cell suspension culture of Daucus carota.

Science.gov (United States)

Miras-Moreno, Begoña; Almagro, Lorena; Pedreño, M A; Sabater-Jara, Ana Belén

2016-09-01

In this work, suspension-cultured cells of Daucus carota were used to evaluate the effect of β-cyclodextrins on the production of isoprenoid and phenolic compounds. The results showed that the phytosterols and phenolic compounds were accumulated in the extracellular medium (15100μgL(-1) and 477.46μgL(-1), respectively) in the presence of cyclodextrins. Unlike the phytosterol and phenolic compound content, β-carotene (1138.03μgL(-1)), lutein (25949.54μgL(-1)) and α-tocopherol (8063.82μgL(-1)) chlorophyll a (1625.13μgL(-1)) and b (9.958 (9958.33μgL(-1)) were mainly accumulated inside the cells. Therefore, cyclodextrins were able to induce the cytosolic mevalonate pathway, increasing the biosynthesis of phytosterols and phenolic compounds, and accumulate them outside the cells. However, in the absence of these cyclic oligosaccharidic elicitors, carrot cells mainly accumulated carotenoids through the methylerythritol 4-phosphate pathway. Therefore, the use of cyclodextrins would allow the extracellular accumulation of both phytosterols and phenolic compounds by diverting the carbon flux towards the cytosolic mevalonate/phenylpropanoid pathway. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Effect of Aluminum Treatment on Proteomes of Radicles of Seeds Derived from Al-Treated Tomato Plants

Directory of Open Access Journals (Sweden)

Ikenna Okekeogbu

2014-03-01

Full Text Available Aluminum (Al toxicity is a major constraint to plant growth and crop yield in acid soils. Tomato cultivars are especially susceptible to excessive Al3+ accumulated in the root zone. In this study, tomato plants were grown in a hydroponic culture system supplemented with 50 µM AlK(SO42. Seeds harvested from Al-treated plants contained a significantly higher Al content than those grown in the control hydroponic solution. In this study, these Al-enriched tomato seeds (harvested from Al-treated tomato plants were germinated in 50 µM AlK(SO42 solution in a homopiperazine-1,4-bis(2-ethanesulfonic acid buffer (pH 4.0, and the control solution which contained the buffer only. Proteomes of radicles were analyzed quantitatively by mass spectrometry employing isobaric tags for relative and absolute quantitation (iTRAQ®. The proteins identified were assigned to molecular functional groups and cellular metabolic pathways using MapMan. Among the proteins whose abundance levels changed significantly were: a number of transcription factors; proteins regulating gene silencing and programmed cell death; proteins in primary and secondary signaling pathways, including phytohormone signaling and proteins for enhancing tolerance to abiotic and biotic stress. Among the metabolic pathways, enzymes in glycolysis and fermentation and sucrolytic pathways were repressed. Secondary metabolic pathways including the mevalonate pathway and lignin biosynthesis were induced. Biological reactions in mitochondria seem to be induced due to an increase in the abundance level of mitochondrial ribosomes and enzymes in the TCA cycle, electron transport chains and ATP synthesis.

Hydrocarbons in the millipede Graphidostreptus tumuliporus (Karsch) (Myriapoda: Diplopoda)—I. In vivo incorporation of 14C-labelled precursors into the hydrocarbon fraction

NARCIS (Netherlands)

Oudejans, R.C.H.M.

1972-01-01

1. 1. Biosynthesis of hydrocarbons in male and female specimens of the millipede Graphidostreptus tumuliporus was investigated after injection of the following precursors: 1-14C-acetate, 16-14C- and 1-14C-palmitic acid, isoleucine-14C(U), valine-14C(U) and mevalonic-2-14C acid. 2. 2. Both sexes

A pilot study of zoledronic acid in the treatment of patients with advanced malignant pleural mesothelioma

Directory of Open Access Journals (Sweden)

Jamil MO

2017-06-01

Full Text Available Muhammad Omer Jamil, Mary S Jerome, Deborah Miley, Katri S Selander, Francisco Robert Division of Hematology and Oncology, Department of Medicine, Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, AL, USA Purpose: Malignant pleural mesothelioma (MPM is a rare malignancy with a dismal median survival of <12 months with current therapy. Single and combination chemotherapy regimens have shown only modest clinical benefit. In preclinical studies, nitrogen-containing bisphosphonates (zoledronic acid inhibit growth of mesothelioma cells by different mechanisms: inhibition of mevalonate pathway, inhibition of angiogenesis, activation of apoptosis through caspase activation, and alteration in activity of matrix metalloproteinases, thereby affecting invasiveness of cancer cells.Patients and methods: We investigated the role of zoledronic acid in a pilot, single-arm trial of MPM patients with Eastern Cooperative Oncology Group (ECOG performance status (PS 0–2 who had progressed on prior treatments or had not received systemic therapy due to poor PS. Primary end point was composite response rate by modified response evaluation criteria in solid tumors and/or metabolic response by 2-deoxy-2-[fluorine-18]fluoro-d-glucose (18F-FDG positron emission tomography criteria. Secondary end points were progression-free survival (PFS and overall survival (OS. Exploratory end points include the effect of zoledronic acid therapy on vascular endothelial growth factor (VEGF, basic fibroblast growth factor, interleukin 8, transforming growth factor beta, mesothelin, and osteopontin levels.Results: Eight male patients (median age of 62 years with the following clinical characteristics were treated; ECOG PS was 0–2, 75% with epithelioid type, and 62% had prior chemotherapy. Overall composite response rate was 12.5% and the clinical benefit rate (response + stable disease was 37.5%. Median PFS was 2 months (0.5–21 months and median OS was

Enhanced volatile fatty acids production from anaerobic fermentation of food waste: A mini-review focusing on acidogenic metabolic pathways.

Science.gov (United States)

Zhou, Miaomiao; Yan, Binghua; Wong, Jonathan W C; Zhang, Yang

2018-01-01

Recently, efficient disposal of food waste (FW) with potential resource recovery has attracted great attentions. Due to its easily biodegradable nature, rich nutrient availability and high moisture content, FW is regarded as favorable substrate for anaerobic digestion (AD). Both waste disposal and energy recovery can be fulfilled during AD of FW. Volatile fatty acids (VFAs) which are the products of the first-two stages of AD, are widely applied in chemical industry as platform chemicals recently. Concentration and distribution of VFAs is the result of acidogenic metabolic pathways, which can be affected by the micro-environment (e.g. pH) in the digester. Hence, the clear elucidation of the acidogenic metabolic pathways is essential for optimization of acidogenic process for efficient product recovery. This review summarizes major acidogenic metabolic pathways and regulating strategies for enhancing VFAs recovery during acidogenic fermentation of FW. Copyright © 2017 Elsevier Ltd. All rights reserved.

Host and Pathway Engineering for Enhanced Lycopene Biosynthesis in Yarrowia lipolytica

Directory of Open Access Journals (Sweden)

Cory Schwartz

2017-11-01

Full Text Available Carotenoids are a class of molecules with commercial value as food and feed additives with nutraceutical properties. Shifting carotenoid synthesis from petrochemical-based precursors to bioproduction from sugars and other biorenewable carbon sources promises to improve process sustainability and economics. In this work, we engineered the oleaginous yeast Yarrowia lipolytica to produce the carotenoid lycopene. To enhance lycopene production, we tested a series of strategies to modify host cell physiology and metabolism, the most successful of which were mevalonate pathway overexpression and alleviating auxotrophies previously engineered into the PO1f strain of Y. lipolytica. The beneficial engineering strategies were combined into a single strain, which was then cultured in a 1-L bioreactor to produce 21.1 mg/g DCW. The optimized strain overexpressed a total of eight genes including two copies of HMG1, two copies of CrtI, and single copies of MVD1, EGR8, CrtB, and CrtE. Recovering leucine and uracil biosynthetic capacity also produced significant enhancement in lycopene titer. The successful engineering strategies characterized in this work represent a significant increase in understanding carotenoid biosynthesis in Y. lipolytica, not only increasing lycopene titer but also informing future studies on carotenoid biosynthesis.

Free fatty acid palmitate activates unfolded protein response pathway and promotes apoptosis in meniscus cells.

Science.gov (United States)

Haywood, J; Yammani, R R

2016-05-01

Obesity is the major risk factor for the development of osteoarthritis (OA); however, the mechanisms involved are not clearly understood. Obesity is associated with increased production of adipokine and elevated levels of circulating free fatty acids (FFA). A recent study has shown that saturated fatty acid palmitate induced pro-inflammatory and pro-apoptotic pathways in chondrocytes. Meniscus has been shown to be more susceptible than articular cartilage to catabolic stimuli. Thus, the aim of this study was to determine the effect of FFA (specifically, palmitate) on meniscus cells. Cultured primary porcine meniscus cells were stimulated with 500 μM FFA (palmitate and oleate) for 24 h to induce endoplasmic reticulum (ER) stress. After treatment, cell lysates were prepared and immunoblotted for C/EBP homologous protein (CHOP). To determine the activation of unfolded protein response (UPR) signaling, cell lysates were probed for cJun n-terminal kinase (JNK), cleaved caspase -3 and Xbp-1s, an alternative mRNA splicing product generated due to Ire1α activation. Treatment of isolated primary meniscus cells with palmitate but not oleate induced expression of CHOP and Xbp-1s. Palmitate treatment of meniscus cells also activated JNK and increased expression of caspase-3, thus promoting apoptosis in meniscus cells. Palmitate induces ER stress and promotes apoptotic pathways in meniscus cells. This is the first study to establish ER stress as a key metabolic mechanistic link between obesity and OA, in addition to (or operating with) biomechanical factors. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

Kynurenine Pathway Metabolites in Humans: Disease and Healthy States

Directory of Open Access Journals (Sweden)

Yiquan Chen

2009-01-01

Full Text Available Tryptophan is an essential amino acid that can be metabolised through different pathways, a major route being the kynurenine pathway. The first enzyme of the pathway, indoleamine-2,3-dioxygenase, is strongly stimulated by inflammatory molecules, particularly interferon gamma. Thus, the kynurenine pathway is often systematically up-regulated when the immune response is activated. The biological significance is that 1 the depletion of tryptophan and generation of kynurenines play a key modulatory role in the immune response; and 2 some of the kynurenines, such as quinolinic acid, 3-hydroxykynurenine and kynurenic acid, are neuroactive. The kynurenine pathway has been demonstrated to be involved in many diseases and disorders, including Alzheimer's disease, amyotrophic lateral sclerosis, Huntington's disease, AIDS dementia complex, malaria, cancer, depression and schizophrenia, where imbalances in tryptophan and kynurenines have been found. This review compiles most of these studies and provides an overview of how the kynurenine pathway might be contributing to disease development, and the concentrations of tryptophan and kynurenines in the serum, cerebrospinal fluid and brain tissues in control and patient subjects.

Engineering a Synthetic Microbial Consortium for Comprehensive Conversion of Algae Biomass into Terpenes for Advanced Biofuels and Bioproducts

Energy Technology Data Exchange (ETDEWEB)

Wu, Weihua [Sandia National Lab. (SNL-CA), Livermore, CA (United States); Wu, Benjamin Chiau-Pin [Sandia National Lab. (SNL-CA), Livermore, CA (United States); Davis, Ryan Wesley [Sandia National Lab. (SNL-CA), Livermore, CA (United States)

2015-10-01

Recent strategies for algae-based biofuels have primarily focused on biodiesel production by exploiting high algal lipid yields under nutrient stress conditions. However, under conditions supporting robust algal biomass accumulation, carbohydrate and proteins typically comprise up to ~80% of the ash-free dry weight of algae biomass. Therefore, comprehensive utilization of algal biomass for production of multipurpose intermediate- to high-value bio-based products will promote scale-up of algae production and processing to commodity volumes. Terpenes are hydrocarbon and hydrocarbon-like (C:O>10:1) compounds with high energy density, and are therefore potentially promising candidates for the next generation of value added bio-based chemicals and “drop-in” replacements for petroleum-based fuels. In this study, we demonstrated the feasibility of bioconversion of proteins into sesquiterpene compounds as well as comprehensive bioconversion of algal carbohydrates and proteins into biofuels. To achieve this, the mevalonate pathway was reconstructed into an E. coli chassis with six different terpene synthases (TSs). Strains containing the various TSs produced a spectrum of sesquiterpene compounds in minimal medium containing amino acids as the sole carbon source. The sesquiterpene production was optimized through three different regulation strategies using chamigrene synthase as an example. The highest total terpene titer reached 166 mg/L, and was achieved by applying a strategy to minimize mevalonate accumulation in vivo. The highest yields of total terpene were produced under reduced IPTG induction levels (0.25 mM), reduced induction temperature (25°C), and elevated substrate concentration (20 g/L amino acid mixture). A synthetic bioconversion consortium consisting of two engineering E. coli strains (DH1-TS and YH40-TS) with reconstructed terpene biosynthetic pathways was designed for comprehensive single-pot conversion of algal carbohydrates and proteins to

Investigating sources and pathways of perfluoroalkyl acids (PFAAs) in aquifers in Tokyo using multiple tracers

International Nuclear Information System (INIS)

Kuroda, Keisuke; Murakami, Michio; Oguma, Kumiko; Takada, Hideshige; Takizawa, Satoshi

2014-01-01

We employed a multi-tracer approach to investigate sources and pathways of perfluoroalkyl acids (PFAAs) in urban groundwater, based on 53 groundwater samples taken from confined aquifers and unconfined aquifers in Tokyo. While the median concentrations of groundwater PFAAs were several ng/L, the maximum concentrations of perfluorooctane sulfonate (PFOS, 990 ng/L), perfluorooctanoate (PFOA, 1800 ng/L) and perfluorononanoate (PFNA, 620 ng/L) in groundwater were several times higher than those of wastewater and street runoff reported in the literature. PFAAs were more frequently detected than sewage tracers (carbamazepine and crotamiton), presumably owing to the higher persistence of PFAAs, the multiple sources of PFAAs beyond sewage (e.g., surface runoff, point sources) and the formation of PFAAs from their precursors. Use of multiple methods of source apportionment including principal component analysis–multiple linear regression (PCA–MLR) and perfluoroalkyl carboxylic acid ratio analysis highlighted sewage and point sources as the primary sources of PFAAs in the most severely polluted groundwater samples, with street runoff being a minor source (44.6% sewage, 45.7% point sources and 9.7% street runoff, by PCA–MLR). Tritium analysis indicated that, while young groundwater (recharged during or after the 1970s, when PFAAs were already in commercial use) in shallow aquifers (< 50 m depth) was naturally highly vulnerable to PFAA pollution, PFAAs were also found in old groundwater (recharged before the 1950s, when PFAAs were not in use) in deep aquifers (50–500 m depth). This study demonstrated the utility of multiple uses of tracers (pharmaceuticals and personal care products; PPCPs, tritium) and source apportionment methods in investigating sources and pathways of PFAAs in multiple aquifer systems. - Highlights: • Aquifers in Tokyo had high levels of perfluoroalkyl acids (up to 1800 ng/L). • PFAAs were more frequently detected than sewage

Investigating sources and pathways of perfluoroalkyl acids (PFAAs) in aquifers in Tokyo using multiple tracers

Energy Technology Data Exchange (ETDEWEB)

Kuroda, Keisuke, E-mail: keisukekr@gmail.com [Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-8656 (Japan); Murakami, Michio [Institute of Industrial Science, The University of Tokyo, 4-6-1 Komaba, Meguro, Tokyo 153-8505 (Japan); Oguma, Kumiko [Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-8656 (Japan); Takada, Hideshige [Laboratory of Organic Geochemistry (LOG), Institute of Symbiotic Science and Technology, Tokyo University of Agriculture and Technology, Fuchu, Tokyo 183-8509 (Japan); Takizawa, Satoshi [Department of Urban Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-8656 (Japan)

2014-08-01

We employed a multi-tracer approach to investigate sources and pathways of perfluoroalkyl acids (PFAAs) in urban groundwater, based on 53 groundwater samples taken from confined aquifers and unconfined aquifers in Tokyo. While the median concentrations of groundwater PFAAs were several ng/L, the maximum concentrations of perfluorooctane sulfonate (PFOS, 990 ng/L), perfluorooctanoate (PFOA, 1800 ng/L) and perfluorononanoate (PFNA, 620 ng/L) in groundwater were several times higher than those of wastewater and street runoff reported in the literature. PFAAs were more frequently detected than sewage tracers (carbamazepine and crotamiton), presumably owing to the higher persistence of PFAAs, the multiple sources of PFAAs beyond sewage (e.g., surface runoff, point sources) and the formation of PFAAs from their precursors. Use of multiple methods of source apportionment including principal component analysis–multiple linear regression (PCA–MLR) and perfluoroalkyl carboxylic acid ratio analysis highlighted sewage and point sources as the primary sources of PFAAs in the most severely polluted groundwater samples, with street runoff being a minor source (44.6% sewage, 45.7% point sources and 9.7% street runoff, by PCA–MLR). Tritium analysis indicated that, while young groundwater (recharged during or after the 1970s, when PFAAs were already in commercial use) in shallow aquifers (< 50 m depth) was naturally highly vulnerable to PFAA pollution, PFAAs were also found in old groundwater (recharged before the 1950s, when PFAAs were not in use) in deep aquifers (50–500 m depth). This study demonstrated the utility of multiple uses of tracers (pharmaceuticals and personal care products; PPCPs, tritium) and source apportionment methods in investigating sources and pathways of PFAAs in multiple aquifer systems. - Highlights: • Aquifers in Tokyo had high levels of perfluoroalkyl acids (up to 1800 ng/L). • PFAAs were more frequently detected than sewage

Enterobacter sp. I-3, a bio-herbicide inhibits gibberellins biosynthetic pathway and regulates abscisic acid and amino acids synthesis to control plant growth.

Science.gov (United States)

Radhakrishnan, Ramalingam; Park, Jae-Man; Lee, In-Jung

2016-12-01

Very few bacterial species were identified as bio-herbicides for weed control. The present research was focused to elucidate the plant growth retardant properties of Enterobacter sp. I-3 during their interaction by determining the changes in endogenous photosynthetic pigments, plant hormones and amino acids. The two bacterial isolates I-4-5 and I-3 were used to select the superior bacterium for controlling weed seeds (Echinochloa crus-galli L. and Portulaca oleracea L.) germination. The post-inoculation of I-3 (Enterobacter sp. I-3) significantly inhibited the weeds seed germination than their controls. The mechanism of bacterium induced plant growth reduction was identified in lettuce treated with I-3 bacterium and compared their effects with known chemical herbicide, trinexapac-ethyl (TE). The treatment of I-3 and TE showed a significant inhibitory effect on shoot length, leaf number, leaf length, leaf width, shoot weight, root weight and chlorophyll content in lettuce seedlings. The endogenous gibberellins (GAs) and abscisic acid (ABA) analysis showed that Enterobacter sp. I-3 treated plants had lower levels of GAs (GA 12 , GA 19 , GA 20 and GA 8 ) and GAs/ABA ratio and then, the higher level of ABA when compared to their controls. Indeed, the individual amino acids ie., aspartic acid, glutamic acid, glycine, threonine, alanine, serine, leucine, isoleucine and tyrosine were declined in TE and I-3 exposed plants. Our results suggest that the utilization of Enterobacter sp. I-3 inhibits the GAs pathway and amino acids synthesis in weeds to control their growth can be an alternative to chemical herbicides. Copyright © 2016 Elsevier GmbH. All rights reserved.

Statins: pros and cons.

Science.gov (United States)

Pinal-Fernandez, Iago; Casal-Dominguez, Maria; Mammen, Andrew L

2018-05-23

Statins inhibit the critical step of cholesterol synthesis in which 3-hydroxy-3-methylglutaryl coenzyme A (HMGC) is transformed to mevalonate by the enzyme HMGC reductase. By doing so, they have a potent lipid-lowering effect that reduces cardiovascular risk and decreases mortality. Since the mevalonate pathway also influences endothelial function, the inflammatory response, and coagulation, the effects of statins reach well beyond their cholesterol lowering properties. As with all drugs, statins may have adverse effects; these include musculoskeletal symptoms, increased risk of diabetes, and higher rates of hemorrhagic stroke. However, the frequency of adverse effects is extremely low and, in selected patient populations, the benefits of statins considerably outweigh the potential risks. Published by Elsevier España, S.L.U.

Biosynthetic origin of the isoprene units in chromenes of Piper aduncum (Piperaceae)

International Nuclear Information System (INIS)

Leite, Ana C.; Lopes, Adriana A.; Bolzani, Vanderlan da S.; Furlan, Maysa; Kato, Massuo J.

2007-01-01

Metabolic studies involving the incorporation of [1- 13 C]-D-glucose into intact leaves of Piper aduncum (Piperaceae) have indicated that both the mevalonate (MVA) and the pyruvate-triose (MEP) non-mevalonate pathways are implicated in the biosynthesis of isoprene moieties present in methyl 2,2-dimethyl-2H-1-chromene-6-carboxylate (1) and methyl 2,2-dimethyl-8-(3'-methyl- 2'-butenyl)-2H-1-chromene-6-carboxylate (2). The pattern of incorporation of label from [1- 13 C]-D-glucose into these chromenes was determined by quantitative 13 C NMR spectroscopy. The results confirmed that biosynthetic compartment of 1 and 2 could either be the plastid and/ or the cytosol or, possibly, an additional compartment such as the plastid inter-membrane space. (author)

The C-Terminal O-S Acyl Shift Pathway under Acidic Condition to Propose Peptide-Thioesters

Directory of Open Access Journals (Sweden)

Bo Mi Kim

2016-11-01

Full Text Available Peptide-thioester is a pivotal intermediate for peptide ligation and N-, C-terminal cyclization. In this study, desired pathway and the side products of two C-terminal handles, hydroxyethylthiol (HET and hydroxypropylthiol (HPT are described in different conditions as well as kinetic studies. In addition, a new mechanism of C-terminal residue racemization is proposed on the basis of differentiation of products derived from the two C-terminal handles in preparing peptide thioesters through an acid-catalyzed tandem thiol switch, first by an intramolecular O-S acyl shift, and then by an intermolecular S-S exchange.

Non-genomic actions of retinoic acid induce pi3k signaling pathway and phosphorylation of nuclear proteins

OpenAIRE

Laserna Mendieta, Emilio J.; Masiá, Susana; Barettino, Domingo

2007-01-01

Retinoic acid (RA), the active form of vitamin A, induces neuroblastoma cells SH-SY5Y to differentiate. In addition to its classical transcriptional actions regulating the expression of specific genes, RA acts in an extra-genomic way, modulating the activity of relevant signalling cascades. In particular, RA treatment of SH-SY5Y neuroblastoma cells results in activation of phosphatidyl-inositol-3-kinase (PI3K) signaling pathway, and this activation is required for RA-induced differentiation (...

Citric acid induces cell-cycle arrest and apoptosis of human immortalized keratinocyte cell line (HaCaT) via caspase- and mitochondrial-dependent signaling pathways.

Science.gov (United States)

Ying, Tsung-Ho; Chen, Chia-Wei; Hsiao, Yu-Ping; Hung, Sung-Jen; Chung, Jing-Gung; Yang, Jen-Hung

2013-10-01

Citric acid is an alpha-hydroxyacid (AHA) widely used in cosmetic dermatology and skincare products. However, there is concern regarding its safety for the skin. In this study, we investigated the cytotoxic effects of citric acid on the human keratinocyte cell line HaCaT. HaCaT cells were treated with citric acid at 2.5-12.5 mM for different time periods. Cell-cycle arrest and apoptosis were investigated by 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) staining, flow cytometry, western blot and confocal microscopy. Citric acid not only inhibited proliferation of HaCaT cells in a dose-dependent manner, but also induced apoptosis and cell cycle-arrest at the G2/M phase (before 24 h) and S phase (after 24 h). Citric acid increased the level of Bcl-2-associated X protein (BAX) and reduced the levels of B-cell lymphoma-2 (BCL-2), B-cell lymphoma-extra large (BCL-XL) and activated caspase-9 and caspase-3, which subsequently induced apoptosis via caspase-dependent and caspase-independent pathways. Citric acid also activated death receptors and increased the levels of caspase-8, activated BH3 interacting-domain death agonist (BID) protein, Apoptosis-inducing factor (AIF), and Endonuclease G (EndoG). Therefore, citric acid induces apoptosis through the mitochondrial pathway in the human keratinocyte cell line HaCaT. The study results suggest that citric acid is cytotoxic to HaCaT cells via induction of apoptosis and cell-cycle arrest in vitro.

Identification and functional characterisation of genes encoding the omega-3 polyunsaturated fatty acid biosynthetic pathway from the coccolithophore Emiliania huxleyi.

Science.gov (United States)

Sayanova, Olga; Haslam, Richard P; Calerón, Monica Venegas; López, Noemi Ruiz; Worthy, Charlotte; Rooks, Paul; Allen, Michael J; Napier, Johnathan A

2011-05-01

The Prymnesiophyceae coccolithophore Emiliania huxleyi is one of the most abundant alga in our oceans and therefore plays a central role in marine foodwebs. E. huxleyi is notable for the synthesis and accumulation of the omega-3 long chain polyunsaturated fatty acid docosahexaenoic acid (DHA; 22:6Δ(4,7,10,13,16,19), n-3) which is accumulated in fish oils and known to have health-beneficial properties to humans, preventing cardiovascular disease and related pathologies. Here we describe the identification and functional characterisation of the five E. huxleyi genes which direct the synthesis of docosahexaenoic acid in this alga. Surprisingly, E. huxleyi does not use the conventional Δ6-pathway, instead using the alternative Δ8-desaturation route which has previously only been observed in a few unrelated microorganisms. Given that E. huxleyi accumulates significant levels of the Δ6-desaturated fatty acid stearidonic acid (18:4Δ(6,9,12,15), n-3), we infer that the biosynthesis of DHA is likely to be metabolically compartmentalised from the synthesis of stearidonic acid. Copyright © 2011 Elsevier Ltd. All rights reserved.

Pentose pathway in human liver

International Nuclear Information System (INIS)

Magnusson, I.; Chandramouli, V.; Schumann, W.C.; Kumaran, K.; Wahren, J.; Landau, B.R.

1988-01-01

[1- 14 C]Ribose and [1- 14 C]glucose were given to normal subjects along with glucose loads (1 g per kg of body weight) after administration of diflunisal and acetaminophen, drugs that are excreted in urine as glucuronides. Distributions of 14 C were determined in the carbons of the excreted glucoronides and in the glucose from blood samples drawn from hepatic veins before and after glucagon administration. Eighty percent or more of the 14 C from [1- 14 C]ribose incorporated into the glucuronic acid moiety of the glucuronides was in carbons 1 and 3, with less than 8% in carbon 2. In glucuronic acid from glucuronide excreted when [2- 14 C]glucose was given, 3.5-8.1% of the 14 C was in carbon 1, 2.5-4.3% in carbon 3, and more than 70% in carbon 2. These distributions are in accord with the glucuronides sampling the glucose unit of the glucose 6-phosphate pool that is a component of the pentose pathway and is intermediate in glycogen formation. It is concluded that the glucuronic acid conjugates of the drugs can serve as a noninvasive means of sampling hepatic glucose 6-phosphate. In human liver, as in animal liver, the classical pentose pathway functions, not the L-type pathway, and only a small percentage of the glucose is metabolized via the pathway

Application of a controllable degron strategy for metabolic engineering

DEFF Research Database (Denmark)

Knuf, Christoph; Maury, Jerome; Jacobsen, Simo Abdessamad

2014-01-01

In numerous cases of metabolic engineering, metabolite pools have to be increased in order to obtain flux into heterologous pathways. A simple tool for this would be the deletion of genes that would practically lead to a block of the natural pathway, so that the carbon can flow into the heterolog...... of intermediates of the mevalonate pathway around 2,3-oxidosqualene, which is the precursor for triterpenoids. Many triterpenoids are pharmaceutically relevant compounds which nowadays need to be extracted from plant material through an intricate and resource consuming process....

In response to 'Can sugars be produced from fatty acids? A test case for pathway analysis tools'.

Science.gov (United States)

Faust, Karoline; Croes, Didier; van Helden, Jacques

2009-12-01

In their article entitled 'Can sugars be produced from fatty acids? A test case for pathway analysis tools' de Figueiredo and co-authors assess the performance of three pathway prediction tools (METATOOL, PathFinding and Pathway Hunter Tool) using the synthesis of glucose-6-phosphate (G6P) from acetyl-CoA in humans as a test case. We think that this article is biased for three reasons: (i) the metabolic networks used as input for the respective tools were of very different sizes; (ii) the 'assessment' is restricted to two study cases; (iii) developers are inherently more skilled to use their own tools than those developed by other people. We extended the analyses led by de Figueiredo and clearly show that the apparent superior performance of their tool (METATOOL) is partly due to the differences in input network sizes. We also see a conceptual problem in the comparison of tools that serve different purposes. In our opinion, metabolic path finding and elementary mode analysis are answering different biological questions, and should be considered as complementary rather than competitive approaches. Supplementary data are available at Bioinformatics online.

Response of tobacco to the Pseudomonas syringae pv. Tomato DC3000 is mainly dependent on salicylic acid signaling pathway.

Science.gov (United States)

Liu, Yang; Wang, Li; Cai, Guohua; Jiang, Shanshan; Sun, Liping; Li, Dequan

2013-07-01

Pseudomonas syringae pv. Tomato DC3000 (Pst DC3000) was the first pathogen to be demonstrated to infect Arabidopsis and to cause disease symptoms in the laboratory setting. However, the defense response to Pst DC3000 was unclear in tobacco. In this report, the expression profiles of twelve defense response-related genes were analyzed after treatment with salicylic acid (SA), jasmonic acid (JA), and pathogen Pst DC3000 by qRT-PCR. According to our results, it could be presented that the genes primarily induced by SA were also induced to higher levels after Pst DC3000 infection. SA accumulation could be induced to a higher level than that of JA after Pst DC3000 infection. In addition, SA could result in hypersensitive response (HR), which did not completely depend on accumulation of reactive oxygen species. These results indicated that tobacco mainly depended on SA signaling pathway rather than on JA signaling pathway in response to Pst DC3000. Further study demonstrated that JA could significantly inhibit the accumulation of SA and the generation of the HR induced by Pst DC3000. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

T3SS-dependent differential modulations of the jasmonic acid pathway in susceptible and resistant genotypes of Malus spp. challenged with Erwinia amylovora.

Science.gov (United States)

Dugé De Bernonville, Thomas; Gaucher, Matthieu; Flors, Victor; Gaillard, Sylvain; Paulin, Jean-Pierre; Dat, James F; Brisset, Marie-Noëlle

2012-06-01

Fire blight is a bacterial disease of Maloideae caused by Erwinia amylovora (Ea). This necrogenic enterobacterium uses a type III secretion system (T3SS) to inject type III effectors into the plant cells to cause disease on its susceptible hosts, including economically important crops like apple and pear. The expressions of marker genes of the salicylic acid (SA) and jasmonic acid (JA) defense regulation pathways were monitored by RT-qPCR in leaves of two apple genotypes, one susceptible and one resistant, challenged with a wild type strain, a T3SS-deficient strain or water. The transcriptional data taken together with hormone level measurements indicated that the SA pathway was similarly induced in both apple genotypes during infection by Ea. On the contrary, the data clearly showed a strong T3SS-dependent down-regulation of the JA pathway in leaves of the susceptible genotype but not in those of the resistant one. Accordingly, methyl-jasmonate treated susceptible plants displayed an increased resistance to Ea. Bacterial mutant analysis indicated that JA manipulation by Ea mainly relies on the type III effector DspA/E. Taken together, our data suggest that the T3SS-dependent down-regulation of the JA pathway is a critical step in the infection process of Malus spp. by Ea. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Metabolic reconstructions identify plant 3-methylglutaconyl-CoA hydratase that is crucial for branched-chain amino acid catabolism in mitochondria.

Science.gov (United States)

Latimer, Scott; Li, Yubing; Nguyen, Thuong T H; Soubeyrand, Eric; Fatihi, Abdelhak; Elowsky, Christian G; Block, Anna; Pichersky, Eran; Basset, Gilles J

2018-05-09

The proteinogenic branched-chain amino acids (BCAAs) leucine, isoleucine and valine are essential nutrients for mammals. In plants, BCAAs double as alternative energy sources when carbohydrates become limiting, the catabolism of BCAAs providing electrons to the respiratory chain and intermediates to the tricarboxylic acid cycle. Yet, the actual architecture of the degradation pathways of BCAAs is not well understood. In this study, gene network modeling in Arabidopsis and rice, and plant-prokaryote comparative genomics detected candidates for 3-methylglutaconyl-CoA hydratase (4.2.1.18), one of the missing plant enzymes of leucine catabolism. Alignments of these protein candidates sampled from various spermatophytes revealed non-homologous N-terminal extensions that are lacking in their bacterial counterparts, and green fluorescent protein-fusion experiments demonstrated that the Arabidopsis protein, product of gene At4g16800, is targeted to mitochondria. Recombinant At4g16800 catalyzed the dehydration of 3-hydroxymethylglutaryl-CoA into 3-methylglutaconyl-CoA, and displayed kinetic features similar to those of its prokaryotic homolog. When at4g16800 knockout plants were subjected to dark-induced carbon starvation, their rosette leaves displayed accelerated senescence as compared to control plants, and this phenotype was paralleled by a marked increase in the accumulation of free and total leucine, isoleucine and valine. The seeds of the at4g16800 mutant showed a similar accumulation of free BCAAs. These data suggest that 3-methylglutaconyl-CoA hydratase is not solely involved in the degradation of leucine, but is also a significant contributor to that of isoleucine and valine. Furthermore, evidence is shown that unlike the situation observed in Trypanosomatidae, leucine catabolism does not contribute to the formation of the terpenoid precursor mevalonate. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights

Trans-Fatty Acids Aggravate Obesity, Insulin Resistance and Hepatic Steatosis in C57BL/6 Mice, Possibly by Suppressing the IRS1 Dependent Pathway.

Science.gov (United States)

Zhao, Xiaona; Shen, Cheng; Zhu, Hong; Wang, Cong; Liu, Xiangwei; Sun, Xiaolei; Han, Shasha; Wang, Peng; Dong, Zhen; Ma, Xin; Hu, Kai; Sun, Aijun; Ge, Junbo

2016-05-30

Trans-fatty acid consumption has been reported as a risk factor for metabolic disorders and targeted organ damages. Nonetheless, little is known about the roles and mechanisms of trans-fatty acids in obesity, insulin resistance (IR) and hepatic steatosis. Adult C57BL/6 male mice were fed with four different diets for 20 weeks: normal diet (ND), high fat diet (HFD), low trans-fatty acids diet (LTD) and high trans-fatty acid diet (HTD). The diet-induced metabolic disorders were assessed by evaluating body weight, glucose tolerance test, hepatic steatosis and plasma lipid profiles post 20-week diet. Histological (H&E, Oil-Red-O) staining and western blot analysis were employed to assess liver steatosis and potential signaling pathways. After 20-weeks of diet, the body weights of the four groups were 29.61 ± 1.89 g (ND), 39.04 ± 4.27 g (HFD), 34.09 ± 2.62 g (LTD) and 43.78 ± 4.27 g (HTD) (p steatosis compared with HFD group possibly through regulating adipose triglyceride lipase. The group consuming the HTD also exhibited significantly reduced levels of IRS1, phosphor-PKC and phosphor-AKT. These results support our hypothesis that consumption of a diet high in trans-fatty acids induces higher rates of obesity, IR and hepatic steatosis in male C57BL/6 mice, possibly by suppressing the IRS1dependent pathway.

The octadecanoid signalling pathway in plants mediates a response to ultraviolet radiation

International Nuclear Information System (INIS)

Conconi, A.; Smerdon, M.J.; Howe, G.A.; Ryan, C.A.

1996-01-01

Many plant genes that respond to environmental and developmental changes are regulated by jasmonic acid, which is derived from linolenic acid via the octadecanoid pathway. Linolenic acid is an important fatty-acid constituent of membranes in most plant species and its intracellular levels increase in response to certain signals. Here we report that irradiation of tomato leaves with ultraviolet light induces the expression of several plant defensive genes that are normally activated through the octadecanoid pathway after wounding. The response to ultraviolet light is blocked by an inhibitor of the octadecanoid pathway and it does not occur in a tomato mutant defective in this pathway. The ultraviolet irradiation maximally induces the defence genes at levels where cyclobutane pyrimidine dimer formation, an indicator of DNA damage, is less than 0.2 dimers per gene. Our evidence indicates that this plant defence response to certain wavelengths of ultraviolet radiation requires the activation of the octadecanoid defence signalling pathway. (author)

Metabolic engineering of the omega-3 long chain polyunsaturated fatty acid biosynthetic pathway into transgenic plants.

Science.gov (United States)

Ruiz-López, Noemi; Sayanova, Olga; Napier, Johnathan A; Haslam, Richard P

2012-04-01

Omega-3 (ω-3) very long chain polyunsaturated fatty acids (VLC-PUFAs) such as eicosapentaenoic acid (EPA; 20:5 Δ5,8,11,14,17) and docosahexaenoic acid (DHA; 22:6 Δ4,7,10,13,16,19) have been shown to have significant roles in human health. Currently the primary dietary source of these fatty acids are marine fish; however, the increasing demand for fish and fish oil (in particular the expansion of the aquaculture industry) is placing enormous pressure on diminishing marine stocks. Such overfishing and concerns related to pollution in the marine environment have directed research towards the development of a viable alternative sustainable source of VLC-PUFAs. As a result, the last decade has seen many genes encoding the primary VLC-PUFA biosynthetic activities identified and characterized. This has allowed the reconstitution of the VLC-PUFA biosynthetic pathway in oilseed crops, producing transgenic plants engineered to accumulate ω-3 VLC-PUFAs at levels approaching those found in native marine organisms. Moreover, as a result of these engineering activities, knowledge of the fundamental processes surrounding acyl exchange and lipid remodelling has progressed. The application of new technologies, for example lipidomics and next-generation sequencing, is providing a better understanding of seed oil biosynthesis and opportunities for increasing the production of unusual fatty acids. Certainly, it is now possible to modify the composition of plant oils successfully, and, in this review, the most recent developments in this field and the challenges of producing VLC-PUFAs in the seed oil of higher plants will be described.

Salicylic Acid and Jasmonic Acid Pathways are Activated in Spatially Different Domains Around the Infection Site During Effector-Triggered Immunity in Arabidopsis thaliana.

Science.gov (United States)

Betsuyaku, Shigeyuki; Katou, Shinpei; Takebayashi, Yumiko; Sakakibara, Hitoshi; Nomura, Nobuhiko; Fukuda, Hiroo

2018-01-01

The innate immune response is, in the first place, elicited at the site of infection. Thus, the host response can be different among the infected cells and the cells surrounding them. Effector-triggered immunity (ETI), a form of innate immunity in plants, is triggered by specific recognition between pathogen effectors and their corresponding plant cytosolic immune receptors, resulting in rapid localized cell death known as hypersensitive response (HR). HR cell death is usually limited to a few cells at the infection site, and is surrounded by a few layers of cells massively expressing defense genes such as Pathogenesis-Related Gene 1 (PR1). This virtually concentric pattern of the cellular responses in ETI is proposed to be regulated by a concentration gradient of salicylic acid (SA), a phytohormone accumulated around the infection site. Recent studies demonstrated that jasmonic acid (JA), another phytohormone known to be mutually antagonistic to SA in many cases, is also accumulated in and required for ETI, suggesting that ETI is a unique case. However, the molecular basis for this uniqueness remained largely to be solved. Here, we found that, using intravital time-lapse imaging, the JA signaling pathway is activated in the cells surrounding the central SA-active cells around the infection sites in Arabidopsis thaliana. This distinct spatial organization explains how these two phythormone pathways in a mutually antagonistic relationship can be activated simultaneously during ETI. Our results re-emphasize that the spatial consideration is a key strategy to gain mechanistic insights into the apparently complex signaling cross-talk in immunity. © The Author 2017. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Ursolic acid inhibits adipogenesis in 3T3-L1 adipocytes through LKB1/AMPK pathway.

Directory of Open Access Journals (Sweden)

Yonghan He

Full Text Available BACKGROUND: Ursolic acid (UA is a triterpenoid compound with multiple biological functions. This compound has recently been reported to possess an anti-obesity effect; however, the mechanisms are less understood. OBJECTIVE: As adipogenesis plays a critical role in obesity, the present study was conducted to investigate the effect of UA on adipogenesis and mechanisms of action in 3T3-L1 preadipocytes. METHODS AND RESULTS: The 3T3-L1 preadipocytes were induced to differentiate in the presence or absence of UA for 6 days. The cells were determined for proliferation, differentiation, fat accumulation as well as the protein expressions of molecular targets that regulate or are involved in fatty acid synthesis and oxidation. The results demonstrated that ursolic acid at concentrations ranging from 2.5 µM to 10 µM dose-dependently attenuated adipogenesis, accompanied by reduced protein expression of CCAAT element binding protein β (C/EBPβ, peroxisome proliferator-activated receptor γ (PPARγ, CCAAT element binding protein α (C/EBPα and sterol regulatory element binding protein 1c (SREBP-1c, respectively. Ursolic acid increased the phosphorylation of acetyl-CoA carboxylase (ACC and protein expression of carnitine palmitoyltransferase 1 (CPT1, but decreased protein expression of fatty acid synthase (FAS and fatty acid-binding protein 4 (FABP4. Ursolic acid increased the phosphorylation of AMP-activated protein kinase (AMPK and protein expression of (silent mating type information regulation 2, homolog 1 (Sirt1. Further studies demonstrated that the anti-adipogenic effect of UA was reversed by the AMPK siRNA, but not by the Sirt1 inhibitor nicotinamide. Liver kinase B1 (LKB1, the upstream kinase of AMPK, was upregulated by UA. When LKB1 was silenced with siRNA or the inhibitor radicicol, the effect of UA on AMPK activation was diminished. CONCLUSIONS: Ursolic acid inhibited 3T3-L1 preadipocyte differentiation and adipogenesis through the LKB1/AMPK

Biosynthetic origin of the isoprene units in chromenes of Piper aduncum (Piperaceae)

Energy Technology Data Exchange (ETDEWEB)

Leite, Ana C.; Lopes, Adriana A.; Bolzani, Vanderlan da S.; Furlan, Maysa [UNESP, Araraquara, SP (Brazil). Inst. de Quimica]. E-mail: maysaf@iq.unesp.br; Kato, Massuo J. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica

2007-07-01

Metabolic studies involving the incorporation of [1-{sup 13}C]-D-glucose into intact leaves of Piper aduncum (Piperaceae) have indicated that both the mevalonate (MVA) and the pyruvate-triose (MEP) non-mevalonate pathways are implicated in the biosynthesis of isoprene moieties present in methyl 2,2-dimethyl-2H-1-chromene-6-carboxylate (1) and methyl 2,2-dimethyl-8-(3'-methyl- 2'-butenyl)-2H-1-chromene-6-carboxylate (2). The pattern of incorporation of label from [1- {sup 13}C]-D-glucose into these chromenes was determined by quantitative {sup 13}C NMR spectroscopy. The results confirmed that biosynthetic compartment of 1 and 2 could either be the plastid and/ or the cytosol or, possibly, an additional compartment such as the plastid inter-membrane space. (author)

WikiPathways: a multifaceted pathway database bridging metabolomics to other omics research.

Science.gov (United States)

Slenter, Denise N; Kutmon, Martina; Hanspers, Kristina; Riutta, Anders; Windsor, Jacob; Nunes, Nuno; Mélius, Jonathan; Cirillo, Elisa; Coort, Susan L; Digles, Daniela; Ehrhart, Friederike; Giesbertz, Pieter; Kalafati, Marianthi; Martens, Marvin; Miller, Ryan; Nishida, Kozo; Rieswijk, Linda; Waagmeester, Andra; Eijssen, Lars M T; Evelo, Chris T; Pico, Alexander R; Willighagen, Egon L

2018-01-04

WikiPathways (wikipathways.org) captures the collective knowledge represented in biological pathways. By providing a database in a curated, machine readable way, omics data analysis and visualization is enabled. WikiPathways and other pathway databases are used to analyze experimental data by research groups in many fields. Due to the open and collaborative nature of the WikiPathways platform, our content keeps growing and is getting more accurate, making WikiPathways a reliable and rich pathway database. Previously, however, the focus was primarily on genes and proteins, leaving many metabolites with only limited annotation. Recent curation efforts focused on improving the annotation of metabolism and metabolic pathways by associating unmapped metabolites with database identifiers and providing more detailed interaction knowledge. Here, we report the outcomes of the continued growth and curation efforts, such as a doubling of the number of annotated metabolite nodes in WikiPathways. Furthermore, we introduce an OpenAPI documentation of our web services and the FAIR (Findable, Accessible, Interoperable and Reusable) annotation of resources to increase the interoperability of the knowledge encoded in these pathways and experimental omics data. New search options, monthly downloads, more links to metabolite databases, and new portals make pathway knowledge more effortlessly accessible to individual researchers and research communities. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Flower abscission in Vitis vinifera L. triggered by gibberellic acid and shade discloses differences in the underlying metabolic pathways

Directory of Open Access Journals (Sweden)

Sara eDomingos

2015-06-01

Full Text Available Understanding abscission is both a biological and an agronomic challenge. Flower abscission induced independently by shade and gibberellic acid (GAc sprays was monitored in grapevine (Vitis vinifera L. growing under a soilless greenhouse system during two seasonal growing conditions, in an early and late production cycle. Physiological and metabolic changes triggered by each of the two distinct stimuli were determined. Environmental conditions exerted a significant effect on fruit set as showed by the higher natural drop rate recorded in the late production cycle with respect to the early cycle. Shade and GAc treatments increased the percentage of flower drop compared to the control, and at a similar degree, during the late production cycle. The reduction of leaf gas exchanges under shade conditions was not observed in GAc treated vines. The metabolic profile assessed in samples collected during the late cycle differently affected primary and secondary metabolisms and showed that most of the treatment-resulting variations occurred in opposite trends in inflorescences unbalanced in either hormonal or energy deficit abscission-inducing signals. Particularly concerning carbohydrates metabolism, sucrose, glucose, tricarboxylic acid (TCA metabolites and intermediates of the raffinose family oligosaccharides pathway were lower in shaded and higher in GAc samples. Altered oxidative stress remediation mechanisms and indolacetic acid (IAA concentration were identified as abscission signatures common to both stimuli. According to the global analysis performed, we report that grape flower abscission mechanisms triggered by GAc application and C-starvation are not based on the same metabolic pathways.

TMV-Cg Coat Protein stabilizes DELLA proteins and in turn negatively modulates salicylic acid-mediated defense pathway during Arabidopsis thaliana viral infection.

Science.gov (United States)

Rodriguez, Maria Cecilia; Conti, Gabriela; Zavallo, Diego; Manacorda, Carlos Augusto; Asurmendi, Sebastian

2014-08-03

Plant viral infections disturb defense regulatory networks during tissue invasion. Emerging evidence demonstrates that a significant proportion of these alterations are mediated by hormone imbalances. Although the DELLA proteins have been reported to be central players in hormone cross-talk, their role in the modulation of hormone signaling during virus infections remains unknown. This work revealed that TMV-Cg coat protein (CgCP) suppresses the salicylic acid (SA) signaling pathway without altering defense hormone SA or jasmonic acid (JA) levels in Arabidopsis thaliana. Furthermore, it was observed that the expression of CgCP reduces plant growth and delays the timing of floral transition. Quantitative RT-qPCR analysis of DELLA target genes showed that CgCP alters relative expression of several target genes, indicating that the DELLA proteins mediate transcriptional changes produced by CgCP expression. Analyses by fluorescence confocal microscopy showed that CgCP stabilizes DELLA proteins accumulation in the presence of gibberellic acid (GA) and that the DELLA proteins are also stabilized during TMV-Cg virus infections. Moreover, DELLA proteins negatively modulated defense transcript profiles during TMV-Cg infection. As a result, TMV-Cg accumulation was significantly reduced in the quadruple-DELLA mutant Arabidopsis plants compared to wild type plants. Taken together, these results demonstrate that CgCP negatively regulates the salicylic acid-mediated defense pathway by stabilizing the DELLA proteins during Arabidopsis thaliana viral infection, suggesting that CgCP alters the stability of DELLAs as a mechanism of negative modulation of antiviral defense responses.

Quantitative evaluation of the biosynthetic pathways leading to δ-aminolevulinic acid from the Shemin precursor glycine via the C5 pathway in Arthrobacter hyalinus by analysis of 13C-labeled coproporphyrinogen III biosynthesized from [2-13C]glycine, [1-13C]acetate, and [2-13C]acetate using 13C NMR spectroscopy

International Nuclear Information System (INIS)

Katsumi Iida

2013-01-01

The biosynthetic pathways leading to δ-aminolevulinic acid (ALA) from the Shemin precursor glycine via the C5 pathway in Arthrobacter hyalinus were quantitatively evaluated by means of feeding experiments with [2- 13 C]glycine, sodium [1- 13 C]acetate, and sodium [2- 13 C]acetate, followed by analysis of the labeling patterns of coproporphyrinogen III (Copro'gen III) (biosynthesized from ALA) using 13 C NMR spectroscopy. Two biosynthetic pathways leading to ALA from glycine via the C5 pathway were identified: i.e., transformation of glycine to l-serine catalyzed by glycine hydroxymethyltransferase, and glycine synthase-catalyzed catabolism of glycine to N 5 , N 10 -methylene-tetrahydrofolic acid (THF), which reacts with another molecule of glycine to afford l-serine. l-Serine is transformed to acetyl-CoA via pyruvic acid. Acetyl-CoA enters the tricarboxylic acid cycle, affording 2-oxoglutaric acid, which in turn is transformed to l-glutamic acid. The l-glutamic acid enters the C5 pathway, affording ALA in A. hyalinus. A 13 C NMR spectroscopic comparison of the labeling patterns of Copro'gen III obtained after feeding of [2- 13 C]glycine, sodium [1- 13 C]acetate, and sodium [2- 13 C]acetate showed that [2- 13 C]glycine transformation and [2- 13 C]glycine catabolism in A. hyalinus proceed in the ratio of 52 and 48 %. The reaction of [2- 13 C]glycine and N 5 , N 10 -methylene-THF, that of glycine and N 5 , N 10 -[methylene- 13 C]methylene-THF generated from the [2- 13 C]glycine catabolism, and that of [2- 13 C]glycine and N 5 , N 10 -[methylene- 13 C]methylene-THF transformed the fed [2- 13 C]glycine to [1- 13 C]acetyl-CoA, [2- 13 C]acetyl-CoA, and [1,2- 13 C 2 ]acetyl-CoA in the ratios of 42, 37, and 21 %, respectively. These labeled acetyl-CoAs were then incorporated into ALA. Our results provide a quantitative picture of the pathways of biosynthetic transformation to ALA from glycine in A. hyalinus. (author)

Simvastatin attenuates acrolein-induced mucin production in rats: involvement of the Ras/extracellular signal-regulated kinase pathway.

Science.gov (United States)

Chen, Ya-Juan; Chen, Peng; Wang, Hai-Xia; Wang, Tao; Chen, Lei; Wang, Xun; Sun, Bei-Bei; Liu, Dai-Shun; Xu, Dan; An, Jing; Wen, Fu-Qiang

2010-06-01

Airway mucus overproduction is a cardinal feature of airway inflammatory diseases, such as chronic obstructive pulmonary disease and cystic fibrosis. Since the small G-protein Ras is known to modulate cellular functions in the lung, we sought to investigate whether the Ras inhibitor simvastatin could attenuate acrolein-induced mucin production in rat airways. Rats were exposed to acrolein for 12 days, after first being pretreated intragastrically for 24 h with either simvastatin alone or simvastatin in combination with mevalonate, which prevents the isoprenylation needed for Ras activation. Lung tissue was analyzed for extracellular signal-regulated kinase (ERK) activity, goblet cell metaplasia and mucin production. To analyze the effect of simvastatin on mucin production in more detail, acrolein-exposed human airway epithelial NCI-H292 cells were pretreated with simvastatin alone or together with mevalonate. Culture medium was collected to detect mucin secretion, and cell lysates were examined for Ras-GTPase activity and epidermal growth factor receptor (EGFR)/ERK phosphorylation. In vivo, simvastatin treatment dose-dependently suppressed acrolein-induced goblet cell hyperplasia and metaplasia in bronchial epithelium and inhibited ERK phosphorylation in rat lung homogenates. Moreover, simvastatin inhibited Muc5AC mucin synthesis at both the mRNA and protein levels in the lung. In vitro, simvastatin pretreatment attenuated the acrolein-induced significant increase in MUC5AC mucin expression, Ras-GTPase activity and EGFR/ERK phosphorylation. These inhibitory effects of simvastatin were neutralized by mevalonate administration both in vitro and in vivo. Our results suggest that simvastatin may attenuate acrolein-induced mucin protein synthesis in the airway and airway inflammation, possibly by blocking ERK activation mediated by Ras protein isoprenylation. Thus, the evidence from the experiment suggests that human trials are warranted to determine the potential

The SUD1 gene encodes a putative E3 ubiquitin ligase and is a positive regulator of 3-hydroxy-3-methylglutaryl coenzyme a reductase activity in Arabidopsis.

Science.gov (United States)

Doblas, Verónica G; Amorim-Silva, Vítor; Posé, David; Rosado, Abel; Esteban, Alicia; Arró, Montserrat; Azevedo, Herlander; Bombarely, Aureliano; Borsani, Omar; Valpuesta, Victoriano; Ferrer, Albert; Tavares, Rui M; Botella, Miguel A

2013-02-01

The 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) enzyme catalyzes the major rate-limiting step of the mevalonic acid (MVA) pathway from which sterols and other isoprenoids are synthesized. In contrast with our extensive knowledge of the regulation of HMGR in yeast and animals, little is known about this process in plants. To identify regulatory components of the MVA pathway in plants, we performed a genetic screen for second-site suppressor mutations of the Arabidopsis thaliana highly drought-sensitive drought hypersensitive2 (dry2) mutant that shows decreased squalene epoxidase activity. We show that mutations in SUPPRESSOR OF DRY2 DEFECTS1 (SUD1) gene recover most developmental defects in dry2 through changes in HMGR activity. SUD1 encodes a putative E3 ubiquitin ligase that shows sequence and structural similarity to yeast Degradation of α factor (Doα10) and human TEB4, components of the endoplasmic reticulum-associated degradation C (ERAD-C) pathway. While in yeast and animals, the alternative ERAD-L/ERAD-M pathway regulates HMGR activity by controlling protein stability, SUD1 regulates HMGR activity without apparent changes in protein content. These results highlight similarities, as well as important mechanistic differences, among the components involved in HMGR regulation in plants, yeast, and animals.

Perfluorononanoic acid-induced apoptosis in rat spleen involves oxidative stress and the activation of caspase-independent death pathway

International Nuclear Information System (INIS)

Fang, Xuemei; Feng, Yixing; Wang, Jianshe; Dai, Jiayin

2010-01-01

Perfluoroalkyl acid (PFAA)-induced apoptosis has been reported in many cell types. However, minimal information on its mode of action is available. This study explored the possible involvement of apoptotic signaling pathways in a nine-carbon-chain length PFAA-perfluorononanoic acid (PFNA)-induced splenocyte apoptosis. After a 14-day exposure to PFNA, rat spleens showed dose-dependent levels of apoptosis. The production of pro-inflammatory and anti-inflammatory cytokines was significantly increased and decreased, respectively. However, protein levels of tumor necrosis factor receptor 1 (TNFR1), fas-associated protein with death domain (FADD), caspase 8 and caspase 3, which are involved in inflammation-related and caspase-dependent apoptosis, were discordant. Peroxisome proliferator-activated receptors alpha (PPARα) and PPARγ genes expression was up-regulated in rats treated with 3 or 5 mg/kg/day of PFNA, and the level of hydrogen peroxide (H 2 O 2 ) increased concurrently in rats treated with the highest dose. Moreover, superoxide dismutase (SOD) activity and Bcl-2 protein levels were dramatically decreased in spleens after treatment with 3 and 5 mg/kg/day of PFNA. However, protein levels of Bax were unchanged. Apoptosis-inducing factor (AIF), an initiator of caspase-independent apoptosis, was significantly increased in all PFNA-dosed rats. Thus, oxidative stress and the activation of a caspase-independent apoptotic signaling pathway contributed to PFNA-induced apoptosis in rat splenocytes.

Oleanolic acid supplement attenuates liquid fructose-induced adipose tissue insulin resistance through the insulin receptor substrate-1/phosphatidylinositol 3-kinase/Akt signaling pathway in rats

Energy Technology Data Exchange (ETDEWEB)

Li, Ying [Faculty of Basic Medical Sciences, Chongqing Medical University, Chongqing 400016 (China); Wang, Jianwei, E-mail: wangjianwei1968@gmail.com [Department of Traditional Chinese Medicine, Chongqing Medical University, Chongqing 400016 (China); Gu, Tieguang [Endocrinology and Metabolism Group, Sydney Institute of Health Sciences, Sydney, NSW 2000 Australia (Australia); Yamahara, Johji [Pharmafood Institute, Kyoto 602-8136 (Japan); Li, Yuhao, E-mail: yuhao@sitcm.edu.au [Endocrinology and Metabolism Group, Sydney Institute of Health Sciences, Sydney, NSW 2000 Australia (Australia)

2014-06-01

Oleanolic acid, a triterpenoid contained in more than 1620 plants including various fruits and foodstuffs, has numerous metabolic effects, such as hepatoprotection. However, its underlying mechanisms remain poorly understood. Adipose tissue insulin resistance (Adipo-IR) may contribute to the development and progress of metabolic abnormalities through release of excessive free fatty acids from adipose tissue. This study investigated the effect of oleanolic acid on Adipo-IR. The results showed that supplement with oleanolic acid (25 mg/kg, once daily, by oral gavage) over 10 weeks attenuated liquid fructose-induced increase in plasma insulin concentration and the homeostasis model assessment of insulin resistance (HOMA-IR) index in rats. Simultaneously, oleanolic acid reversed the increase in the Adipo-IR index and plasma non-esterified fatty acid concentrations during the oral glucose tolerance test assessment. In white adipose tissue, oleanolic acid enhanced mRNA expression of the genes encoding insulin receptor, insulin receptor substrate (IRS)-1 and phosphatidylinositol 3-kinase. At the protein level, oleanolic acid upregulated total IRS-1 expression, suppressed the increased phosphorylated IRS-1 at serine-307, and restored the increased phosphorylated IRS-1 to total IRS-1 ratio. In contrast, phosphorylated Akt to total Akt ratio was increased. Furthermore, oleanolic acid reversed fructose-induced decrease in phosphorylated-Akt/Akt protein to plasma insulin concentration ratio. However, oleanolic acid did not affect IRS-2 mRNA expression. Therefore, these results suggest that oleanolic acid supplement ameliorates fructose-induced Adipo-IR in rats via the IRS-1/phosphatidylinositol 3-kinase/Akt pathway. Our findings may provide new insights into the mechanisms of metabolic actions of oleanolic acid. - Highlights: • Adipose insulin resistance (Adipo-IR) contributes to metabolic abnormalities. • We investigated the effect of oleanolic acid (OA) on adipo-IR in

Oleanolic acid supplement attenuates liquid fructose-induced adipose tissue insulin resistance through the insulin receptor substrate-1/phosphatidylinositol 3-kinase/Akt signaling pathway in rats

International Nuclear Information System (INIS)

Li, Ying; Wang, Jianwei; Gu, Tieguang; Yamahara, Johji; Li, Yuhao

2014-01-01

Oleanolic acid, a triterpenoid contained in more than 1620 plants including various fruits and foodstuffs, has numerous metabolic effects, such as hepatoprotection. However, its underlying mechanisms remain poorly understood. Adipose tissue insulin resistance (Adipo-IR) may contribute to the development and progress of metabolic abnormalities through release of excessive free fatty acids from adipose tissue. This study investigated the effect of oleanolic acid on Adipo-IR. The results showed that supplement with oleanolic acid (25 mg/kg, once daily, by oral gavage) over 10 weeks attenuated liquid fructose-induced increase in plasma insulin concentration and the homeostasis model assessment of insulin resistance (HOMA-IR) index in rats. Simultaneously, oleanolic acid reversed the increase in the Adipo-IR index and plasma non-esterified fatty acid concentrations during the oral glucose tolerance test assessment. In white adipose tissue, oleanolic acid enhanced mRNA expression of the genes encoding insulin receptor, insulin receptor substrate (IRS)-1 and phosphatidylinositol 3-kinase. At the protein level, oleanolic acid upregulated total IRS-1 expression, suppressed the increased phosphorylated IRS-1 at serine-307, and restored the increased phosphorylated IRS-1 to total IRS-1 ratio. In contrast, phosphorylated Akt to total Akt ratio was increased. Furthermore, oleanolic acid reversed fructose-induced decrease in phosphorylated-Akt/Akt protein to plasma insulin concentration ratio. However, oleanolic acid did not affect IRS-2 mRNA expression. Therefore, these results suggest that oleanolic acid supplement ameliorates fructose-induced Adipo-IR in rats via the IRS-1/phosphatidylinositol 3-kinase/Akt pathway. Our findings may provide new insights into the mechanisms of metabolic actions of oleanolic acid. - Highlights: • Adipose insulin resistance (Adipo-IR) contributes to metabolic abnormalities. • We investigated the effect of oleanolic acid (OA) on adipo-IR in

Regulatory cross-talks and cascades in rice hormone biosynthesis pathways contribute to stress signaling

Directory of Open Access Journals (Sweden)

Arindam Deb

2016-08-01

Full Text Available Crosstalk among different hormone signaling pathways play an important role in modulating plant response to both biotic and abiotic stress. Hormone activity is controlled by its bio-availability, which is again influenced by its biosynthesis. Thus independent hormone biosynthesis pathways must be regulated and co-ordinated to mount an integrated response. One of the possibilities is to use cis-regulatory elements to orchestrate expression of hormone biosynthesis genes. Analysis of CREs, associated with differentially expressed hormone biosynthesis related genes in rice leaf under Magnaporthe oryzae attack and drought stress enabled us to obtain insights about cross-talk among hormone biosynthesis pathways at the transcriptional level. We identified some master transcription regulators that co-ordinate different hormone biosynthesis pathways under stress. We found that Abscisic acid and Brassinosteroid regulate Cytokinin conjugation; conversely Brassinosteroid biosynthesis is affected by both Abscisic acid and Cytokinin. Jasmonic acid and Ethylene biosynthesis may be modulated by Abscisic acid through DREB transcription factors. Jasmonic acid or Salicylic acid biosynthesis pathways are co-regulated but they are unlikely to influence each other’s production directly. Thus multiple hormones may modulate hormone biosynthesis pathways through a complex regulatory network, where biosynthesis of one hormone is affected by several other contributing hormones.

Improving monoterpene geraniol production through geranyl diphosphate synthesis regulation in Saccharomyces cerevisiae.

Science.gov (United States)

Zhao, Jianzhi; Bao, Xiaoming; Li, Chen; Shen, Yu; Hou, Jin

2016-05-01

Monoterpenes have wide applications in the food, cosmetics, and medicine industries and have recently received increased attention as advanced biofuels. However, compared with sesquiterpenes, monoterpene production is still lagging in Saccharomyces cerevisiae. In this study, geraniol, a valuable acyclic monoterpene alcohol, was synthesized in S. cerevisiae. We evaluated three geraniol synthases in S. cerevisiae, and the geraniol synthase Valeriana officinalis (tVoGES), which lacked a plastid-targeting peptide, yielded the highest geraniol production. To improve geraniol production, synthesis of the precursor geranyl diphosphate (GPP) was regulated by comparing three specific GPP synthase genes derived from different plants and the endogenous farnesyl diphosphate synthase gene variants ERG20 (G) (ERG20 (K197G) ) and ERG20 (WW) (ERG20 (F96W-N127W) ), and controlling endogenous ERG20 expression, coupled with increasing the expression of the mevalonate pathway by co-overexpressing IDI1, tHMG1, and UPC2-1. The results showed that overexpressing ERG20 (WW) and strengthening the mevalonate pathway significantly improved geraniol production, while expressing heterologous GPP synthase genes or down-regulating endogenous ERG20 expression did not show positive effect. In addition, we constructed an Erg20p(F96W-N127W)-tVoGES fusion protein, and geraniol production reached 66.2 mg/L after optimizing the amino acid linker and the order of the proteins. The best strain yielded 293 mg/L geraniol in a fed-batch cultivation, a sevenfold improvement over the highest titer previously reported in an engineered S. cerevisiae strain. Finally, we showed that the toxicity of geraniol limited its production. The platform developed here can be readily used to synthesize other monoterpenes.

Development of an image-based screening system for inhibitors of the plastidial MEP pathway and of protein geranylgeranylation

Science.gov (United States)

Hartmann, Michael; Gas-Pascual, Elisabet; Hemmerlin, Andrea; Rohmer, Michel; Bach, Thomas J.

2015-01-01

In a preceding study we have recently established an in vivo visualization system for the geranylgeranylation of proteins in a stably transformed tobacco BY-2 cell line, which involves expressing a dexamethasone-inducible GFP fused to the prenylable, carboxy-terminal basic domain of the rice calmodulin CaM61, which naturally bears a CaaL geranylgeranylation motif (GFP-BD-CVIL). By using pathway-specific inhibitors it was there demonstrated that inhibition of the methylerythritol phosphate (MEP) pathway with oxoclomazone and fosmidomycin, as well as inhibition of protein geranylgeranyl transferase type 1 (PGGT-1), shifted the localization of the GFP-BD-CVIL protein from the membrane to the nucleus. In contrast, the inhibition of the mevalonate (MVA) pathway with mevinolin did not affect this localization. Furthermore, in this initial study complementation assays with pathway-specific intermediates confirmed that the precursors for the cytosolic isoprenylation of this fusion protein are predominantly provided by the MEP pathway. In order to optimize this visualization system from a more qualitative assay to a statistically trustable medium or a high-throughput screening system, we established now new conditions that permit culture and analysis in 96-well microtiter plates, followed by fluorescence microscopy. For further refinement, the existing GFP-BD-CVIL cell line was transformed with an estradiol-inducible vector driving the expression of a RFP protein, C-terminally fused to a nuclear localization signal (NLS-RFP). We are thus able to quantify the total number of viable cells versus the number of inhibited cells after various treatments. This approach also includes a semi-automatic counting system, based on the freely available image processing software. As a result, the time of image analysis as well as the risk of user-generated bias is reduced to a minimum. Moreover, there is no cross-induction of gene expression by dexamethasone and estradiol, which is an

Rewiring Host Lipid Metabolism by Large Viruses Determines the Fate of Emiliania huxleyi, a Bloom-Forming Alga in the Ocean[C][W][OPEN

Science.gov (United States)

Rosenwasser, Shilo; Mausz, Michaela A.; Schatz, Daniella; Sheyn, Uri; Malitsky, Sergey; Aharoni, Asaph; Weinstock, Eyal; Tzfadia, Oren; Ben-Dor, Shifra; Feldmesser, Ester; Pohnert, Georg; Vardi, Assaf

2014-01-01

Marine viruses are major ecological and evolutionary drivers of microbial food webs regulating the fate of carbon in the ocean. We combined transcriptomic and metabolomic analyses to explore the cellular pathways mediating the interaction between the bloom-forming coccolithophore Emiliania huxleyi and its specific coccolithoviruses (E. huxleyi virus [EhV]). We show that EhV induces profound transcriptome remodeling targeted toward fatty acid synthesis to support viral assembly. A metabolic shift toward production of viral-derived sphingolipids was detected during infection and coincided with downregulation of host de novo sphingolipid genes and induction of the viral-encoded homologous pathway. The depletion of host-specific sterols during lytic infection and their detection in purified virions revealed their novel role in viral life cycle. We identify an essential function of the mevalonate-isoprenoid branch of sterol biosynthesis during infection and propose its downregulation as an antiviral mechanism. We demonstrate how viral replication depends on the hijacking of host lipid metabolism during the chemical “arms race” in the ocean. PMID:24920329

Jasmonate-induced biosynthesis of andrographolide in Andrographis paniculata.

Science.gov (United States)

Sharma, Shiv Narayan; Jha, Zenu; Sinha, Rakesh Kumar; Geda, Arvind Kumar

2015-02-01

Andrographolide is a prominent secondary metabolite found in Andrographis paniculata that exhibits enormous pharmacological effects. In spite of immense value, the normal biosynthesis of andrographolide results in low amount of the metabolite. To induce the biosynthesis of andrographolide, we attempted elicitor-induced activation of andrographolide biosynthesis in cell cultures of A. paniculata. This was carried out by using methyl jasmonate (MeJA) as an elicitor. Among the various concentrations of MeJA tested at different time periods, 5 µM MeJA yielded 5.25 times more andrographolide content after 24 h of treatment. The accumulation of andrographolide was correlated with the expression level of known regulatory genes (hmgs, hmgr, dxs, dxr, isph and ggps) of mevalonic acid (MVA) and 2-C-methyl-d-erythritol-4-phosphate (MEP) pathways. These results established the involvement of MeJA in andrographolide biosynthesis by inducing the transcription of its biosynthetic pathways genes. The coordination of isph, ggps and hmgs expression highly influenced the andrographolide biosynthesis. © 2014 Scandinavian Plant Physiology Society.

Linoleic acid enhance the production of moncolin K and red pigments in Monascus ruber by activating mokH and mokA, and by accelerating cAMP-PkA pathway.

Science.gov (United States)

Huang, Jing; Liao, NanQing; Li, HaoMing

2018-04-01

Monacolin K, an inhibitor of HMG-CoA reductase, is a secondary metabolite synthesized by polyketide synthases (PKS) from Monascus ruber. The mokH gene encoding Zn(II)2Cys6 binding protein and mokA gene encoding polyketide synthase are presumed to activate monacolin K production. In this study, linoleic acid could be a quorum sensing signaling molecule to increase monacolin K production in the cyclic AMP(cAMP)-protein kinase A(PKA) signaling pathway. Analysis of the PKA activity and the cAMP concentration shows that linoleic acid could increase cAMP concentration and activate PKA. Analysis of the RT-qPCR products demonstrates that 256μM and 512μM linoleic acid can up-regulate mokH and mokA gene transcript levels. Especially with 512μM linoleic acid addition, linoleic acid increase 1.35 folds of monacolin K production, but 64μM linoleic acid increase 1.94 folds of red pigment production in Monascus ruber. These results show the cAMP-PkA pathway activity can up-regulate mokA and mokH gene, which enhance the yield of Monacolin K. Copyright © 2017 Elsevier B.V. All rights reserved.

Characterization of cyanobacterial hydrocarbon composition and distribution of biosynthetic pathways.

Directory of Open Access Journals (Sweden)

R Cameron Coates

Full Text Available Cyanobacteria possess the unique capacity to naturally produce hydrocarbons from fatty acids. Hydrocarbon compositions of thirty-two strains of cyanobacteria were characterized to reveal novel structural features and insights into hydrocarbon biosynthesis in cyanobacteria. This investigation revealed new double bond (2- and 3-heptadecene and methyl group positions (3-, 4- and 5-methylheptadecane for a variety of strains. Additionally, results from this study and literature reports indicate that hydrocarbon production is a universal phenomenon in cyanobacteria. All cyanobacteria possess the capacity to produce hydrocarbons from fatty acids yet not all accomplish this through the same metabolic pathway. One pathway comprises a two-step conversion of fatty acids first to fatty aldehydes and then alkanes that involves a fatty acyl ACP reductase (FAAR and aldehyde deformylating oxygenase (ADO. The second involves a polyketide synthase (PKS pathway that first elongates the acyl chain followed by decarboxylation to produce a terminal alkene (olefin synthase, OLS. Sixty-one strains possessing the FAAR/ADO pathway and twelve strains possessing the OLS pathway were newly identified through bioinformatic analyses. Strains possessing the OLS pathway formed a cohesive phylogenetic clade with the exception of three Moorea strains and Leptolyngbya sp. PCC 6406 which may have acquired the OLS pathway via horizontal gene transfer. Hydrocarbon pathways were identified in one-hundred-forty-two strains of cyanobacteria over a broad phylogenetic range and there were no instances where both the FAAR/ADO and the OLS pathways were found together in the same genome, suggesting an unknown selective pressure maintains one or the other pathway, but not both.

Relevance of the palatal protein kinase A pathway to the pathogenesis of cleft palate by secalonic acid D in mice

International Nuclear Information System (INIS)

Dhulipala, Vamsidhara C.; Hanumegowda, Umesh M.; Balasubramanian, Ganesh; Reddy, Chada S.

2004-01-01

Secalonic acid-D (SAD) is a teratogenic mycotoxin inducing cleft palate (CP) in the offspring of the exposed mice by reducing palatal shelf size secondary to reduced proliferation of the palatal mesenchymal (PM) cells. Co-administration of dimethylsulfoxide (DMSO) reversed the CP-inducing effect of SAD. Although SAD has been shown to affect both protein kinases A (PKA) and C (PKC) pathways, the relevance of each of these pathways to its CP induction is unknown. The present studies were designed to test the hypothesis that the protective effect of DMSO is mediated by its specific reversal of the effect(s) of SAD on one of these two pathways using ELISA-based activity assays, Western blot analysis, electrophoretic mobility shift assays (EMSA), and murine embryonic PM (MEPM) cell growth in culture. Within the PKA pathway, SAD inhibited the activity of the catalytic subunit of PKA and its migration into the nucleus, elevated phosphorylated cyclic AMP (cAMP) response element (CRE)-binding protein (pCREB) level, and reduced the binding of CREB to CRE. In the PKC pathway, SAD reduced the activity of PKC and the binding of transcription factors (TF) to 12-O-tetradecanoate-13 phorbol acetate-response element (TRE). SAD also inhibited MEPM cell growth and the expression of the CRE- and TRE-containing gene, proliferating cell nuclear antigen (PCNA). Reversal, by DMSO, of the effects of SAD on MEPM cell growth, on PCNA expression and on all components of the PKA, but not of PKC, pathway suggests that the perturbation of the PKA pathway by SAD is relevant to its induction of CP in mice

Pre-steady-state kinetic analysis of 1-deoxy-D-xylulose-5-phosphate reductoisomerase from Mycobacterium tuberculosis reveals partially rate-limiting product release by parallel pathways.

Science.gov (United States)

Liu, Juan; Murkin, Andrew S

2012-07-03

As part of the non-mevalonate pathway for the biosynthesis of the isoprenoid precursor isopentenyl pyrophosphate, 1-deoxy-D-xylulose-5-phosphate (DXP) reductoisomerase (DXR) catalyzes the conversion of DXP into 2-C-methyl-D-erythritol 4-phosphate (MEP) by consecutive isomerization and NADPH-dependent reduction reactions. Because this pathway is essential to many infectious organisms but is absent in humans, DXR is a target for drug discovery. In an attempt to characterize its kinetic mechanism and identify rate-limiting steps, we present the first complete transient kinetic investigation of DXR. Stopped-flow fluorescence measurements with Mycobacterium tuberculosis DXR (MtDXR) revealed that NADPH and MEP bind to the free enzyme and that the two bind together to generate a nonproductive ternary complex. Unlike the Escherichia coli orthologue, MtDXR exhibited a burst in the oxidation of NADPH during pre-steady-state reactions, indicating a partially rate-limiting step follows chemistry. By monitoring NADPH fluorescence during these experiments, the transient generation of MtDXR·NADPH·MEP was observed. Global kinetic analysis supports a model involving random substrate binding and ordered release of NADP(+) followed by MEP. The partially rate-limiting release of MEP occurs via two pathways--directly from the binary complex and indirectly via the MtDXR·NADPH·MEP complex--the partitioning being dependent on NADPH concentration. Previous mechanistic studies, including kinetic isotope effects and product inhibition, are discussed in light of this kinetic mechanism.

Simvastatin Inhibits Goblet Cell Hyperplasia and Lung Arginase in a Mouse Model of Allergic Asthma: A Novel Treatment for Airway Remodeling?

Science.gov (United States)

Zeki, Amir A.; Bratt, Jennifer M.; Rabowsky, Michelle; Last, Jerold A.; Kenyon, Nicholas J.

2010-01-01

Airway remodeling in asthma contributes to airway hyperreactivity, loss of lung function, and persistent symptoms. Current therapies do not adequately treat the structural airway changes associated with asthma. The statins are cholesterol-lowering drugs that inhibit the enzyme 3-hydroxy-3-methyl-glutaryl-CoA reductase, the rate-limiting step of cholesterol biosynthesis in the mevalonate pathway. These drugs have been associated with improved respiratory health and ongoing clinical trials are testing their therapeutic potential in asthma. We hypothesized that simvastatin treatment of ovalbumin-exposed mice would attenuate early features of airway remodeling, by a mevalonate-dependent mechanism. BALB/c mice were initially sensitized to ovalbumin, and then exposed to 1% ovalbumin aerosol for 2 weeks after sensitization for a total of six exposures. Simvastatin (40 mg/kg) or simvastatin plus mevalonate (20 mg/kg) were injected intraperitoneally before each ovalbumin exposure. Treatment with simvastatin attenuated goblet cell hyperplasia, arginase-1 protein expression, and total arginase enzyme activity, but did not alter airway hydroxyproline content or transforming growth factor-β1. Inhibition of goblet cell hyperplasia by simvastatin was mevalonate-dependent. No appreciable changes to airway smooth muscle cells were observed in any of the control or treatment groups. In conclusion, in an acute mouse model of allergic asthma, simvastatin inhibited early hallmarks of airway remodeling, indicators that can lead to airway thickening and fibrosis. Statins are potentially novel treatments for airway remodeling in asthma. Further studies utilizing sub-chronic or chronic allergen exposure models are needed to extend these initial findings. PMID:21078495

Are pleiotropic effects of statins real?

Directory of Open Access Journals (Sweden)

Alberto Corsini

2007-11-01

Full Text Available Alberto Corsini, Nicola Ferri, Michele CortellaroDepartment of Pharmacological Sciences and Department of Clinical Sciences, “Luigi Sacco”, University of Milan, Milan, ItalyAbstract: The clinical benefits of statins are strongly related to their low density lipoproteincholesterol (LDL-C lowering properties. However, because mevalonic acid (MVA, the product of 3-hydroxy-3-methyl-3-glutaryl coenzyme A (HMG-CoA reductase reaction, is the precursor not only of cholesterol but also of nonsteroidal isoprenoid compounds, the inhibition of HMG-CoA reductase may result in pleiotropic effects, independent of their hypocholesterolemic properties. The discrimination between the pleiotropic from LDL-C lowering effects may potentially be more evident during the early phase of treatment since plasma MVA levels drop up to 70% within 1–2 hours while a reduction of LDL-C, detectable after 24 hours, became significant after 6–7 days. Therefore, the deprivation of circulating MVA-derived isoprenoids in the early phase of treatment could be the main mechanism responsible for the atheroprotective effect of statins. This early window of protection in the absence of LDL-C lowering suggests that the anti-inflammatory and the pleiotropic properties of statins may have clinical importance. Therefore, acute coronary syndromes could represent a clinical condition for addressing the early benefits of statins therapy, ie, within 24 h of the event, independent of LDL-C lowering.Keywords: anti-inflammatory effects of statins, mevalonate pathway, LDL lowering, acute coronary syndrome, prenylated proteins

10-Oxo-trans-11-octadecenoic acid generated from linoleic acid by a gut lactic acid bacterium Lactobacillus plantarum is cytoprotective against oxidative stress

Energy Technology Data Exchange (ETDEWEB)

Furumoto, Hidehiro; Nanthirudjanar, Tharnath [Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan); Kume, Toshiaki; Izumi, Yasuhiko [Department of Pharmacology, Graduate School of Pharmaceutical Sciences, Kyoto University, 46-29, Simoadachi-cho, Sakyo-ku, Kyoto 606-8501 (Japan); Park, Si-Bum [Laboratory of Industrial Microbiology, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan); Kitamura, Nahoko; Kishino, Shigenobu; Ogawa, Jun [Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan); Hirata, Takashi [Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan); Faculty of Rehabilitation, Shijonawategakuen University, 5-11-10, Hojo, Daitou-shi, Osaka 574-0011 (Japan); Sugawara, Tatsuya, E-mail: sugawara@kais.kyoto-u.ac.jp [Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502 (Japan)

2016-04-01

Oxidative stress is a well-known cause of multiple diseases. The nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway plays a central role in cellular antioxidative responses. In this study, we investigated the effects of novel fatty acid metabolite derivatives of linoleic acid generated by the gut lactic acid bacteria Lactobacillus plantarum on the Nrf2-ARE pathway. 10-Oxo-trans-11-octadecenoic acid (KetoC) protected HepG2 cells from cytotoxicity induced by hydrogen peroxide. KetoC also significantly increased cellular Nrf2 protein levels, ARE-dependent transcription, and the gene expression of antioxidative enzymes such as heme oxygenase-1 (HO-1), glutamate-cysteine ligase modifier subunit (GCLM), and NAD(P)H:quinone oxidoreductase 1 (NQO1) in HepG2 cells. Additionally, a single oral dose administration of KetoC also increased antioxidative gene expression and protein levels of Nrf2 and HO-1 in mouse organs. Since other fatty acid metabolites and linoleic acid did not affect cellular antioxidative responses, the cytoprotective effect of KetoC may be because of its α,β-unsaturated carbonyl moiety. Collectively, our data suggested that KetoC activated the Nrf2-ARE pathway to enhance cellular antioxidative responses in vitro and in vivo, which further suggests that KetoC may prevent multiple diseases induced by oxidative stress. - Highlights: • We evaluated the effect of modified fatty acids generated by Lactobacillus plantarum. • 10-Oxo-trans-11-ocatadecenoic acid (KetoC) protected cells from oxidative stress. • KetoC activated the Nrf2-ARE pathway to promote antioxidative gene expression. • KetoC promoted the expression of antioxidative enzymes in mice organs. • The cytoprotective effect of KetoC was because of α,β-unsaturated carbonyl moiety.

10-Oxo-trans-11-octadecenoic acid generated from linoleic acid by a gut lactic acid bacterium Lactobacillus plantarum is cytoprotective against oxidative stress

International Nuclear Information System (INIS)

Furumoto, Hidehiro; Nanthirudjanar, Tharnath; Kume, Toshiaki; Izumi, Yasuhiko; Park, Si-Bum; Kitamura, Nahoko; Kishino, Shigenobu; Ogawa, Jun; Hirata, Takashi; Sugawara, Tatsuya

2016-01-01

Oxidative stress is a well-known cause of multiple diseases. The nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway plays a central role in cellular antioxidative responses. In this study, we investigated the effects of novel fatty acid metabolite derivatives of linoleic acid generated by the gut lactic acid bacteria Lactobacillus plantarum on the Nrf2-ARE pathway. 10-Oxo-trans-11-octadecenoic acid (KetoC) protected HepG2 cells from cytotoxicity induced by hydrogen peroxide. KetoC also significantly increased cellular Nrf2 protein levels, ARE-dependent transcription, and the gene expression of antioxidative enzymes such as heme oxygenase-1 (HO-1), glutamate-cysteine ligase modifier subunit (GCLM), and NAD(P)H:quinone oxidoreductase 1 (NQO1) in HepG2 cells. Additionally, a single oral dose administration of KetoC also increased antioxidative gene expression and protein levels of Nrf2 and HO-1 in mouse organs. Since other fatty acid metabolites and linoleic acid did not affect cellular antioxidative responses, the cytoprotective effect of KetoC may be because of its α,β-unsaturated carbonyl moiety. Collectively, our data suggested that KetoC activated the Nrf2-ARE pathway to enhance cellular antioxidative responses in vitro and in vivo, which further suggests that KetoC may prevent multiple diseases induced by oxidative stress. - Highlights: • We evaluated the effect of modified fatty acids generated by Lactobacillus plantarum. • 10-Oxo-trans-11-ocatadecenoic acid (KetoC) protected cells from oxidative stress. • KetoC activated the Nrf2-ARE pathway to promote antioxidative gene expression. • KetoC promoted the expression of antioxidative enzymes in mice organs. • The cytoprotective effect of KetoC was because of α,β-unsaturated carbonyl moiety.

Health risk assessment of various metal(loid)s via multiple exposure pathways on children living near a typical lead-acid battery plant, China

International Nuclear Information System (INIS)

Cao, Suzhen; Duan, Xiaoli; Zhao, Xiuge; Wang, Beibei; Ma, Jin; Fan, Delong; Sun, Chengye; He, Bin; Wei, Fusheng; Jiang, Guibin

2015-01-01

Manufacture of lead-acid batteries is of widespread interest because of its emissions of heavy metals and metalloids into environment, harming environmental quality and consequently causing detrimental effects on human health. In this study, exposure pathways and health risks of children to heavy metal(loid)s (Pb, Cd, As, etc) were investigated based on field sampling and questionnaire. Pb was one of the most abundant elements in children's blood, with an elevated blood lead level of 12.45 μg dL −1 . Soil/dust and food were heavily polluted by targeted metal(loid)s. Food ingestion accounted for more than 80% of the total exposure for most metal(loid)s. The non-cancer risks to children were 3–10 times higher than the acceptable level of 1, while the cancer risks were 5–200 times higher than the maximum acceptable level of 1.0 × 10 −4 . The study emphasized the significance of effective environmental management, particularly to ensure food security near battery facilities. - Highlights: • The health risks of children living around a typical lead-acid battery was analyzed. • The exposure pathways of children to 12 heavy metal(loid)s were assessed. • Courtyard soil and indoor dust and duplicate food were contaminated by metal(loid)s. • Food ingestion was the major pathway for children's exposure to most metal(loid)s. • Higher potentially non-cancer and cancer risks happened to the local children. - The children living around a typical lead-acid battery plant suffered from serious health risks, which mainly attributed to food ingestion and air inhalation exposure

Bile acid metabolism in cirrhosis. VIII. Quantitative evaluation of bile acid synthesis from [7 beta-3H]7 alpha-hydroxycholesterol and [G-3H]26-hydroxycholesterol

International Nuclear Information System (INIS)

Goldman, M.; Vlahcevic, Z.R.; Schwartz, C.C.; Gustafsson, J.; Swell, L.

1982-01-01

In order to evaluate more definitively the observed aberrations in the synthesis of cholic and chenodeoxycholic acids in patients with advanced cirrhosis, two bile acid biosynthesis pathways were examined by determining the efficiency of conversion of [ 3 H]7 alpha-hydroxycholesterol and [ 3 H] 26-hydroxycholesterol to primary bile acids. Bile acid kinetics were determined by administration of [ 14 C]cholic and [ 14 C]chenodeoxycholic acids. Cholic acid synthesis in cirrhotic patients was markedly depressed (170 vs 927 μmoles per day)( while chenodeoxycholic acid synthesis was reduced to a much lesser degree (227 vs 550 μmoles per day). The administration of [ 3 H]7 alpha-hydroxycholesterol allowed for an evaluation of the major pathway of bile acid synthesis via the 7 alpha-hydroxylation of cholesterol. This compound was efficiently incorporated into primary bile acids by the two normal subjects (88 and 100%) and two cirrhotic patients (77 and 91%). However, the recovery of the label in cholic acid was slightly less in cirrhotic patients than in normal subjects. [ 3 H]26-hydroxycholesterol was administered to ascertain the contribution of the 26-hydroxylation pathway to bile acid synthesis. All study subjects showed poor conversion (9 to 22%) of this intermediate into bile acids. The results of this study suggest that a major block in the bile acid synthesis pathway in cirrhosis is at the level of 7 alpha-hydroxylation of cholesterol (impairment of 7 alpha-hydroxylase) and/or in the feedback triggering mechanism regulating bile acid synthesis. The data also suggest that the 26-hydroxylation pathway in normal subjects and patients with cirrhosis is a minor contributor to synthesis of the primary bile acids. Therefore, the relative sparing of chenodeoxycholic acid synthesis observed in cirrhotic patients is not due to preferential synthesis of this bile acid via the 26-hydroxylation pathway

Ethylene Responses in Rice Roots and Coleoptiles Are Differentially Regulated by a Carotenoid Isomerase-Mediated Abscisic Acid Pathway[OPEN

Science.gov (United States)

Yin, Cui-Cui; Ma, Biao; Collinge, Derek Phillip; Pogson, Barry James; He, Si-Jie; Xiong, Qing; Duan, Kai-Xuan; Chen, Hui; Yang, Chao; Lu, Xiang; Wang, Yi-Qin; Zhang, Wan-Ke; Chu, Cheng-Cai; Sun, Xiao-Hong; Fang, Shuang; Chu, Jin-Fang; Lu, Tie-Gang; Chen, Shou-Yi; Zhang, Jin-Song

2015-01-01

Ethylene and abscisic acid (ABA) act synergistically or antagonistically to regulate plant growth and development. ABA is derived from the carotenoid biosynthesis pathway. Here, we analyzed the interplay among ethylene, carotenoid biogenesis, and ABA in rice (Oryza sativa) using the rice ethylene response mutant mhz5, which displays a reduced ethylene response in roots but an enhanced ethylene response in coleoptiles. We found that MHZ5 encodes a carotenoid isomerase and that the mutation in mhz5 blocks carotenoid biosynthesis, reduces ABA accumulation, and promotes ethylene production in etiolated seedlings. ABA can largely rescue the ethylene response of the mhz5 mutant. Ethylene induces MHZ5 expression, the production of neoxanthin, an ABA biosynthesis precursor, and ABA accumulation in roots. MHZ5 overexpression results in enhanced ethylene sensitivity in roots and reduced ethylene sensitivity in coleoptiles. Mutation or overexpression of MHZ5 also alters the expression of ethylene-responsive genes. Genetic studies revealed that the MHZ5-mediated ABA pathway acts downstream of ethylene signaling to inhibit root growth. The MHZ5-mediated ABA pathway likely acts upstream but negatively regulates ethylene signaling to control coleoptile growth. Our study reveals novel interactions among ethylene, carotenogenesis, and ABA and provides insight into improvements in agronomic traits and adaptive growth through the manipulation of these pathways in rice. PMID:25841037

CD147 reprograms fatty acid metabolism in hepatocellular carcinoma cells through Akt/mTOR/SREBP1c and P38/PPARα pathways.

Science.gov (United States)

Li, Jibin; Huang, Qichao; Long, Xiaoyu; Zhang, Jing; Huang, Xiaojun; Aa, Jiye; Yang, Hushan; Chen, Zhinan; Xing, Jinliang

2015-12-01

CD147 is a transmembrane glycoprotein which is highly expressed in various human cancers including hepatocellular carcinoma (HCC). A drug Licartin developed with (131)Iodine-labeled antibody against CD147 has been approved by the Chinese Food and Drug Administration (FDA) and enters into clinical use for HCC treatment. Increasing lines of evidence indicate that CD147 is implicated in the metabolism of cancer cells, especially glycolysis. However, the molecular mechanism underlying the relationship between CD147 and aberrant tumor lipid metabolism remains elusive. We systematically investigated the role of CD147 in the regulation of lipid metabolism, including de novo lipogenesis and fatty acid β-oxidation, in HCC cells and explored the underlying molecular mechanisms. Bioinformatic analysis and experimental evidence demonstrated that CD147 significantly contributed to the reprogramming of fatty acid metabolism in HCC cells mainly through two mechanisms. On one hand, CD147 upregulated the expression of sterol regulatory element binding protein 1c (SREBP1c) by activating the Akt/mTOR signaling pathway, which in turn directly activated the transcription of major lipogenic genes FASN and ACC1 to promote de novo lipogenesis. On the other hand, CD147 downregulated peroxisome proliferator-activated receptor alpha (PPARα) and its transcriptional target genes CPT1A and ACOX1 by activating the p38 MAPK signaling pathway to inhibit fatty acid β-oxidation. Moreover, in vitro and in vivo assays indicated that the CD147-mediated reprogramming of fatty acid metabolism played a critical role in the proliferation and metastasis of HCC cells. Our findings demonstrate that CD147 is a critical regulator of fatty acid metabolism, which provides a strong line of evidence for this molecule to be used as a drug target in cancer treatment. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Primary Metabolic Pathways and Metabolic Flux Analysis

DEFF Research Database (Denmark)

Villadsen, John

2015-01-01

his chapter introduces the metabolic flux analysis (MFA) or stoichiometry-based MFA, and describes the quantitative basis for MFA. It discusses the catabolic pathways in which free energy is produced to drive the cell-building anabolic pathways. An overview of these primary pathways provides...... the reader who is primarily trained in the engineering sciences with atleast a preliminary introduction to biochemistry and also shows how carbon is drained off the catabolic pathways to provide precursors for cell mass building and sometimes for important industrial products. The primary pathways...... to be examined in the following are: glycolysis, primarily by the EMP pathway, but other glycolytic pathways is also mentioned; fermentative pathways in which the redox generated in the glycolytic reactions are consumed; reactions in the tricarboxylic acid (TCA) cycle, which produce biomass precursors and redox...

Enhancing microbial production of biofuels by expanding microbial metabolic pathways.

Science.gov (United States)

Yu, Ping; Chen, Xingge; Li, Peng

2017-09-01

Fatty acid, isoprenoid, and alcohol pathways have been successfully engineered to produce biofuels. By introducing three genes, atfA, adhE, and pdc, into Escherichia coli to expand fatty acid pathway, up to 1.28 g/L of fatty acid ethyl esters can be achieved. The isoprenoid pathway can be expanded to produce bisabolene with a high titer of 900 mg/L in Saccharomyces cerevisiae. Short- and long-chain alcohols can also be effectively biosynthesized by extending the carbon chain of ketoacids with an engineered "+1" alcohol pathway. Thus, it can be concluded that expanding microbial metabolic pathways has enormous potential for enhancing microbial production of biofuels for future industrial applications. However, some major challenges for microbial production of biofuels should be overcome to compete with traditional fossil fuels: lowering production costs, reducing the time required to construct genetic elements and to increase their predictability and reliability, and creating reusable parts with useful and predictable behavior. To address these challenges, several aspects should be further considered in future: mining and transformation of genetic elements related to metabolic pathways, assembling biofuel elements and coordinating their functions, enhancing the tolerance of host cells to biofuels, and creating modular subpathways that can be easily interconnected. © 2016 International Union of Biochemistry and Molecular Biology, Inc.

A SNARE-protein has opposing functions in penetration resistance and defence signalling pathways

DEFF Research Database (Denmark)

Zhang, Ziguo; Feechan, Angela; Pedersen, Carsten

2007-01-01

Penetration resistance is often the first line of defence against fungal pathogens. Subsequently induced defences are mediated by the programmed cell death (PCD) reaction pathway and the salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) signalling pathways. We previously demonstrated...

Trichloroethylene degradation by two independent aromatic-degrading pathways in Alcaligenes eutrophus JMP134.

OpenAIRE

Harker, A R; Kim, Y

1990-01-01

The bacterium Alcaligenes eutrophus JMP134(pJP4) degrades trichloroethylene (TCE) by a chromosomal phenol-dependent pathway and by the plasmid-encoded 2,4-dichlorophenoxyacetic acid pathway. The two pathways were independent and exhibited different rates of removal and capacities for quantity of TCE removed. The phenol-dependent pathway was more rapid (0.2 versus 0.06 nmol of TCE removed per min per mg of protein) and consumed all detectable TCE. The 2,4-dichlorophenoxyacetic acid-dependent p...

The investment in scent: time-resolved metabolic processes in developing volatile-producing Nigella sativa L. seeds.

Directory of Open Access Journals (Sweden)

Wentao Xue

Full Text Available The interplay of processes in central and specialized metabolisms during seed development of Nigella sativa L. was studied by using a high-throughput metabolomics technology and network-based analysis. Two major metabolic shifts were identified during seed development: the first was characterized by the accumulation of storage lipids (estimated as total fatty acids and N-compounds, and the second by the biosynthesis of volatile organic compounds (VOCs and a 30% average decrease in total fatty acids. Network-based analysis identified coordinated metabolic processes during development and demonstrated the presence of five network communities. Enrichment analysis indicated that different compound classes, such as sugars, amino acids, and fatty acids, are largely separated and over-represented in certain communities. One community displayed several terpenoids and the central metabolites, shikimate derived amino acids, raffinose, xylitol and glycerol-3-phosphate. The latter are related to precursors of the mevalonate-independent pathway for VOC production in the plastid; also plastidial fatty acid 18∶3n-3 abundant in "green" seeds grouped with several major terpenes. The findings highlight the interplay between the components of central metabolism and the VOCs. The developmental regulation of Nigella seed metabolism during seed maturation suggests a substantial re-allocation of carbon from the breakdown of fatty acids and from N-compounds, probably towards the biosynthesis of VOCs.

Redox Signaling and CBF-Responsive Pathway Are Involved in Salicylic Acid-Improved Photosynthesis and Growth under Chilling Stress in Watermelon

Science.gov (United States)

Cheng, Fei; Lu, Junyang; Gao, Min; Shi, Kai; Kong, Qiusheng; Huang, Yuan; Bie, Zhilong

2016-01-01

Salicylic acid (SA) plays an important role in plant response to abiotic stresses. This study investigated the potential role of SA in alleviating the adverse effects of chilling stress on photosynthesis and growth in watermelon (Citrullus lanatus). Chilling stress induced the simultaneous accumulation of free and conjugated SA in watermelon plants, and the chilling-induced SA production was attributed to the phenylalanine ammonia-lyase pathway. Applying SA at moderate concentrations induced chilling tolerance, whereas inhibition of SA biosynthesis by L-α-aminooxy-β-phenylpropionic acid (AOPP) increased the photooxidation of PS II under chilling stress in watermelon, resulting in reduced photosynthesis and growth. Chilling induced a transient increase in the ratios of reduced to oxidized glutathione and reduced ascorbate to dehydroascorbate. Then, the expression of antioxidant genes was upregulated, and the activities of antioxidant enzymes were enhanced. Furthermore, SA-induced chilling tolerance was associated with cellular glutathione and ascorbate homeostasis, which served as redox signals to regulate antioxidant metabolism under chilling stress. AOPP treatment stimulated the chilling-induced expression of cold-responsive genes, particularly via C-repeat binding factors CBF3 and CBF4. These results confirm the synergistic role of SA signaling and the CBF-dependent responsive pathway during chilling stress in watermelon. PMID:27777580

Redox Signaling and CBF-Responsive Pathway are Involved in Salicylic Acid-Improved Photosynthesis and Growth under Chilling Stress in Watermelon

Directory of Open Access Journals (Sweden)

Fei Cheng

2016-10-01

Full Text Available Salicylic acid (SA plays an important role in plant response to abiotic stresses. This study investigated the potential role of SA in alleviating the adverse effects of chilling stress on photosynthesis and growth in watermelon (Citrullus lanatus. Chilling stress induced the simultaneous accumulation of free and conjugated SA in watermelon plants, and the chilling-induced SA production was attributed to the phenylalanine ammonia-lyase pathway. Applying SA at moderate concentrations induced chilling tolerance, whereas inhibition of SA biosynthesis by L-ɑ-aminooxy-β-phenylpropionic acid (AOPP increased the photooxidation of PS II under chilling stress in watermelon, resulting in reduced photosynthesis and growth. Chilling induced a transient increase in the ratios of reduced to oxidized glutathione and reduced ascorbate to dehydroascorbate. Then, the expression of antioxidant genes was upregulated, and the activities of antioxidant enzymes were enhanced. Furthermore, SA-induced chilling tolerance was associated with cellular glutathione and ascorbate homeostasis, which served as redox signals to regulate antioxidant metabolism under chilling stress. AOPP treatment stimulated the chilling-induced expression of cold-responsive genes, particularly via C-repeat binding factors CBF3 and CBF4. These results confirm the synergistic role of SA signaling and the CBF-dependent responsive pathway during chilling stress in watermelon.

The metabolism of phytanic acid and pristanic acid in man: a review

NARCIS (Netherlands)

Verhoeven, N. M.; Wanders, R. J.; Poll-The, B. T.; Saudubray, J. M.; Jakobs, C.

1998-01-01

The branched-chain fatty acid phytanic acid is a constituent of the diet, present in diary products, meat and fish. Degradation of this fatty acid in the human body is preceded by activation to phytanoyl-CoA and starts with one cycle of alpha-oxidation. Intermediates in this pathway are

Simvastatin Inhibits IL-5-Induced Chemotaxis and CCR3 Expression of HL-60-Derived and Human Primary Eosinophils.

Science.gov (United States)

Fu, Chia-Hsiang; Tsai, Wan-Chun; Lee, Ta-Jen; Huang, Chi-Che; Chang, Po-Hung; Su Pang, Jong-Hwei

2016-01-01

IL-5-induced chemotaxis of eosinophils is an important feature of allergic airway inflammatory diseases. Simvastatin, a lipid lowering agent, has been shown to exhibit anti-inflammatory and anti-allergic effects. Our aim was to investigate the effect of simvastatin on IL-5-induced eosinophil chemotaxis and its regulatory mechanisms. Eosinophils were derived by treating HL-60 clone 15 (HC15) cells with butyric acid (BA) in an alkaline condition or through direct isolation from human peripheral blood. The expressions of CC chemokine receptor 3 (CCR3) and interleukin (IL)-5 receptors (IL5Rα and β) were analyzed using RT/real-time PCR. The granular proteins were stained using fast green. Eotaxin-induced chemotaxis was measured using a transwell migration assay. CCR3 protein expression was revealed by immunocytochemistry. An animal model of allergic rhinitis was established by challenging Sprague-Dawley® rats repeatedly with ovalbumin. Butyric acid significantly increased the expression of IL5Rα and IL5Rβ, CCR3 and granular proteins in HC15 cells, indicating the maturation of eosinophils (BA-E cells). IL-5 further enhanced the CCR3 expression at both the mRNA and protein levels and the eotaxin-induced chemotaxis of BA-E cells. Simvastatin inhibited the effects of IL-5 on BA-E cells, but not in the presence of mevalonate. Similar results were also exhibited in human primary eosinophils. In vivo animal studies further confirmed that oral simvastatin could significantly suppress the infiltration of eosinophils into turbinate tissues of allergic rats. Therefore, simvastatin was demonstrated to inhibit IL-5-induced CCR3 expression and chemotaxis of eosinophils mediated via the mevalonate pathway. We confirmed that simvastatin also reduced eosinophilic infiltration in allergic rhinitis.

Ferulic acid attenuates diabetes-induced cognitive impairment in rats via regulation of PTP1B and insulin signaling pathway.

Science.gov (United States)

Wang, Hao; Sun, Xiaoxu; Zhang, Ning; Ji, Zhouye; Ma, Zhanqiang; Fu, Qiang; Qu, Rong; Ma, Shiping

2017-12-01

Cognitive impairment has been recognized as a typical characteristic of neurodegenerative disease in diabetes mellitus (DM) and this cognitive dysfunction may be a risk factor for Alzheimer's disease (AD). Ferulic acid, a phenolic compound commonly found in a range of plants, has emerged various properties including anti-inflammatory and neuroprotective effects. In the present study, the protective activities and relevant mechanisms of ferulic acid were evaluated in diabetic rats with cognitive deficits, which were induced by a high-glucose-fat (HGF) diet and low dose of streptozotocin (STZ). It was observed that ferulic acid significantly increased body weight and decreased blood glucose levels. Meanwhile, ferulic acid could markedly ameliorate spatial memory of diabetic rats in Morris water maze (MWM) and decrease AD-like pathologic changes (Aβ deposition and Tau phosphorylation) in the hippocampus, which might be correlated with the inhibition of inflammatory cytokines release and reduction of protein tyrosine phosphatase 1B (PTP1B) expression. Moreover, the levels of brain insulin signal molecules p-IRS, p-Akt and p-GSK3β were also investigated. We found that ferulic acid administration restored the alterations in insulin signaling. In conclusion, ferulic acid exhibited beneficial effects on diabetes-induced cognition lesions, which was involved in the regulation of PTP1B and insulin signaling pathway. We suppose that PTP1B inhibition may represent a promising approach to correct abnormal signaling linked to diabetes-induced cognitive impairment. Copyright © 2017. Published by Elsevier Inc.

Newborn serum retinoic acid level is associated with variants of genes in the retinol metabolism pathway.

Science.gov (United States)

Manolescu, Daniel C; El-Kares, Reyhan; Lakhal-Chaieb, Lajmi; Montpetit, Alexandre; Bhat, Pangala V; Goodyer, Paul

2010-06-01

Retinoic acid (RA) is a critical regulator of gene expression during embryonic development. In rodents, moderate maternal vitamin A deficiency leads to subtle morphogenetic defects and inactivation of RA pathway genes causes major disturbances of embryogenesis. In this study, we quantified RA in umbilical cord blood of 145 healthy full-term Caucasian infants from Montreal. Sixty seven percent of values were ROL). However, we found that the (A) allele of the rs12591551 single nucleotide polymorphism (SNP) in the ALDH1A2 gene (ALDH1A2rs12591551(A)), occurring in 19% of newborns, was associated with 2.5-fold higher serum RA levels. ALDH1A2 encodes retinaldehyde dehydrogenase (RALDH) 2, which synthesizes RA in fetal tissues. We also found that homozygosity for the (A) allele of the rs12724719 SNP in the CRABP2 gene (CRABP2rs12724719(A/A)) was associated with 4.4-fold increase in umbilical cord serum RA. CRABP2 facilitates RA binding to its cognate receptor complex and transfer to the nucleus. We hypothesize that individual variation in RA pathway genes may account for subtle variations in RA-dependent human embryogenesis.

A nuclear-receptor-dependent phosphatidylcholine pathway with antidiabetic effects.

Science.gov (United States)

Lee, Jae Man; Lee, Yoon Kwang; Mamrosh, Jennifer L; Busby, Scott A; Griffin, Patrick R; Pathak, Manish C; Ortlund, Eric A; Moore, David D

2011-05-25

Nuclear hormone receptors regulate diverse metabolic pathways and the orphan nuclear receptor LRH-1 (also known as NR5A2) regulates bile acid biosynthesis. Structural studies have identified phospholipids as potential LRH-1 ligands, but their functional relevance is unclear. Here we show that an unusual phosphatidylcholine species with two saturated 12 carbon fatty acid acyl side chains (dilauroyl phosphatidylcholine (DLPC)) is an LRH-1 agonist ligand in vitro. DLPC treatment induces bile acid biosynthetic enzymes in mouse liver, increases bile acid levels, and lowers hepatic triglycerides and serum glucose. DLPC treatment also decreases hepatic steatosis and improves glucose homeostasis in two mouse models of insulin resistance. Both the antidiabetic and lipotropic effects are lost in liver-specific Lrh-1 knockouts. These findings identify an LRH-1 dependent phosphatidylcholine signalling pathway that regulates bile acid metabolism and glucose homeostasis.

Molecular evolution of the lysine biosynthetic pathways.

Science.gov (United States)

Velasco, A M; Leguina, J I; Lazcano, A

2002-10-01

Among the different biosynthetic pathways found in extant organisms, lysine biosynthesis is peculiar because it has two different anabolic routes. One is the diaminopimelic acid pathway (DAP), and the other over the a-aminoadipic acid route (AAA). A variant of the AAA route that includes some enzymes involved in arginine and leucine biosyntheses has been recently reported in Thermus thermophilus (Nishida et al. 1999). Here we describe the results of a detailed genomic analysis of each of the sequences involved in the two lysine anabolic routes, as well as of genes from other routes related to them. No evidence was found of an evolutionary relationship between the DAP and AAA enzymes. Our results suggest that the DAP pathway is related to arginine metabolism, since the lysC, asd, dapC, dapE, and lysA genes from lysine biosynthesis are related to the argB, argC, argD, argE, and speAC genes, respectively, whose products catalyze different steps in arginine metabolism. This work supports previous reports on the relationship between AAA gene products and some enzymes involved in leucine biosynthesis and the tricarboxylic acid cycle (Irvin and Bhattacharjee 1998; Miyazaki et al. 2001). Here we discuss the significance of the recent finding that several genes involved in the arginine (Arg) and leucine (Leu) biosynthesis participate in a new alternative route of the AAA pathway (Miyazaki et al. 2001). Our results demonstrate a clear relationship between the DAP and Arg routes, and between the AAA and Leu pathways.

Synthetic protein scaffolds based on peptide motifs and cognate adaptor domains for improving metabolic productivity

Directory of Open Access Journals (Sweden)

Anselm H.C. Horn

2015-11-01

Full Text Available The efficiency of many cellular processes relies on the defined interaction among different proteins within the same metabolic or signaling pathway. Consequently, a spatial colocalization of functionally interacting proteins has frequently emerged during evolution. This concept has been adapted within the synthetic biology community for the purpose of creating artificial scaffolds. A recent advancement of this concept is the use of peptide motifs and their cognate adaptor domains. SH2, SH3, GBD, and PDZ domains have been used most often in research studies to date. The approach has been successfully applied to the synthesis of a variety of target molecules including catechin, D-glucaric acid, H2, hydrochinone, resveratrol, butyrate, gamma-aminobutyric acid, and mevalonate. Increased production levels of up to 77-fold have been observed compared to non-scaffolded systems. A recent extension of this concept is the creation of a covalent linkage between peptide motifs and adaptor domains, which leads to a more stable association of the scaffolded systems and thus bears the potential to further enhance metabolic productivity.

Poly(lactic-co-glycolic) acid drug delivery systems through transdermal pathway: an overview.

Science.gov (United States)

Naves, Lucas; Dhand, Chetna; Almeida, Luis; Rajamani, Lakshminarayanan; Ramakrishna, Seeram; Soares, Graça

2017-05-01

In past few decades, scientists have made tremendous advancement in the field of drug delivery systems (DDS), through transdermal pathway, as the skin represents a ready and large surface area for delivering drugs. Efforts are in progress to design efficient transdermal DDS that support sustained drug release at the targeted area for longer duration in the recommended therapeutic window without producing side-effects. Poly(lactic-co-glycolic acid) (PLGA) is one of the most promising Food and Drug Administration approved synthetic polymers in designing versatile drug delivery carriers for different drug administration routes, including transdermal drug delivery. The present review provides a brief introduction over the transdermal drug delivery and PLGA as a material in context to its role in designing drug delivery vehicles. Attempts are made to compile literatures over PLGA-based drug delivery vehicles, including microneedles, nanoparticles, and nanofibers and their role in transdermal drug delivery of different therapeutic agents. Different nanostructure evaluation techniques with their working principles are briefly explained.

Effects of thyroid hormone status on metabolic pathways of arachidonic acid in mice and humans: A targeted metabolomic approach.

Science.gov (United States)

Yao, Xuan; Sa, Rina; Ye, Cheng; Zhang, Duo; Zhang, Shengjie; Xia, Hongfeng; Wang, Yu-cheng; Jiang, Jingjing; Yin, Huiyong; Ying, Hao

2015-01-01

Symptoms of cardiovascular diseases are frequently found in patients with hypothyroidism and hyperthyroidism. However, it is unknown whether arachidonic acid metabolites, the potent mediators in cardiovascular system, are involved in cardiovascular disorders caused by hyperthyroidism and hypothyroidism. To answer this question, serum levels of arachidonic acid metabolites in human subjects with hypothyroidism, hyperthyroidism and mice with hypothyroidism or thyroid hormone treatment were determined by a mass spectrometry-based method. Over ten arachidonic acid metabolites belonging to three catalytic pathways: cyclooxygenases, lipoxygenases, and cytochrome P450, were quantified simultaneously and displayed characteristic profiles under different thyroid hormone status. The level of 20-hydroxyeicosatetraenoic acid, a cytochrome P450 metabolite, was positively correlated with thyroid hormone level and possibly contributed to the elevated blood pressured in hyperthyroidism. The increased prostanoid (PG) I2 and decreased PGE2 levels in hypothyroid patients might serve to alleviate atherosclerosis associated with dyslipidemia. The elevated level of thromboxane (TX) A2, as indicated by TXB2, in hyperthyroid patients and mice treated with thyroid hormone might bring about pulmonary hypertension frequently found in hyperthyroid patients. In conclusion, our prospective study revealed that arachidonic acid metabolites were differentially affected by thyroid hormone status. Certain metabolites may be involved in cardiovascular disorders associated with thyroid diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

Arginine-dependent acid-resistance pathway in Shigella boydii

Science.gov (United States)

Ability to survive the low pH of the human stomach is considered be an important virulent determinant. Acid tolerance of Shigella boydii 18 CDPH, the strain implicated in an outbreak may have played an important role in surviving the acidic food (bean salad). The strain was capable of inducing arg...

Plasma metabolomics reveal the correlation of metabolic pathways and Prakritis of humans

Directory of Open Access Journals (Sweden)

Amey Shirolkar

2018-04-01

Full Text Available Background: Ayurveda, an ancient Indian medicinal system, has categorized human body constitutions in three broad constitutional types (prakritis i.e. Vata, Pitta and Kapha. Objectives: Analysis of plasma metabolites and related pathways to classify Prakriti specific dominant marker metabolites and metabolic pathways. Materials and methods: 38 healthy male individuals were assessed for dominant Prakritis and their fasting blood samples were collected. The processed plasma samples were subjected to rapid resolution liquid chromatography–electrospray ionization–quadrupole time of flight mass spectrometry (RRLC–ESI–QTOFMS. Mass profiles were aligned and subjected to multivariate analysis. Results: Partial least square discriminant analysis (PLS-DA model showed 97.87% recognition capability. List of PLS-DA metabolites was subjected to permutative Benjamini–Hochberg false discovery rate (FDR correction and final list of 76 metabolites with p  2.0 was identified. Pathway analysis using metascape and JEPETTO plugins in Cytoscape revealed that steroidal hormone biosynthesis, amino acid, and arachidonic acid metabolism are major pathways varying with different constitution. Biological Go processes analysis showed that aromatic amino acids, sphingolipids, and pyrimidine nucleotides metabolic processes were dominant in kapha type of body constitution. Fat soluble vitamins, cellular amino acid, and androgen biosynthesis process along with branched chain amino acid and glycerolipid catabolic processes were dominant in pitta type individuals. Vata Prakriti was found to have dominant catecholamine, arachidonic acid and hydrogen peroxide metabolomics processes. Conclusion: The neurotransmission and oxidative stress in vata, BCAA catabolic, androgen, xenobiotics metabolic processes in pitta, and aromatic amino acids, sphingolipid, and pyrimidine metabolic process in kapha Prakriti were the dominant marker pathways. Keywords: Ayurveda, Prakriti, Human

UV-C-Induced alleviation of transcriptional gene silencing through plant-plant communication: Key roles of jasmonic acid and salicylic acid pathways.

Science.gov (United States)

Xu, Wei; Wang, Ting; Xu, Shaoxin; Li, Fanghua; Deng, Chenguang; Wu, Lijun; Wu, Yuejin; Bian, Po

2016-08-01

Plant stress responses at the epigenetic level are expected to allow more permanent changes of gene expression and potentially long-term adaptation. While it has been reported that plants subjected to adverse environments initiate various stress responses in their neighboring plants, little is known regarding epigenetic responses to external stresses mediated by plant-plant communication. In this study, we show that DNA repetitive elements of Arabidopsis thaliana, whose expression is inhibited epigenetically by transcriptional gene silencing (TGS) mechanism, are activated by UV-C irradiation through airborne plant-plant and plant-plant-plant communications, accompanied by DNA demethylation at CHH sites. Moreover, the TGS is alleviated by direct treatments with exogenous methyl jasmonate (MeJA) and methyl salicylate (MeSA). Further, the plant-plant and plant-plant-plant communications are blocked by mutations in the biosynthesis or signaling of jasmonic acid (JA) or salicylic acid (SA), indicating that JA and SA pathways are involved in the interplant communication for epigenetic responses. For the plant-plant-plant communication, stress cues are relayed to the last set of receiver plants by promoting the production of JA and SA signals in relaying plants, which exhibit upregulated expression of genes for JA and SA biosynthesis and enhanced emanation of MeJA and MeSA. Copyright © 2016 Elsevier B.V. All rights reserved.

Characterization and regulation of Leishmania major 3-hydroxy-3-methylglutaryl-CoA reductase

DEFF Research Database (Denmark)

Montalvetti, A; Pena Diaz, Javier; Hurtado, R

2000-01-01

In eukaryotes the enzyme 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) reductase catalyses the synthesis of mevalonic acid, a common precursor to all isoprenoid compounds. Here we report the isolation and overexpression of the gene coding for HMG-CoA reductase from Leishmania major. The protein from L...

Bardoxolone methyl (BARD) ameliorates aristolochic acid (AA)-induced acute kidney injury through Nrf2 pathway.

Science.gov (United States)

Wu, Juan; Liu, Xinhui; Fan, Jinjin; Chen, Wenfang; Wang, Juan; Zeng, Youjia; Feng, Xiaorang; Yu, Xueqing; Yang, Xiao

2014-04-06

Bardoxolone methyl (BARD) is an antioxidant modulator that acts through induction of the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. This study aimed to investigate the role of BARD in protecting kidneys from aristolochic acid (AA)-induced acute kidney injury (AKI). Male C57BL/6 mice received intraperitoneal (i.p.) injections of aristolochic acid I (AAI) (5mg/kg/day) for 5 days to produce acute AA nephropathy (AAN) model. BARD (10mg/kg/day, i.p.) was applied for 7 consecutive days, starting 2 days prior to AAI administration. The mice in the AA group showed AKI as evidenced by worsening kidney function evaluated by blood urea nitrogen (BUN) and serum creatinine (SCr) levels, and severe tubulointerstitial injury marked by massive tubule necrosis in kidney tissues. BARD significantly reduced BUN and SCr levels which were elevated by AAI. Additionally, AAI-induced histopathological renal damage was ameliorated by BARD. Furthermore, the expression of Nrf2 was reduced, and its repressor Kelch-like ECH-associated protein 1 (Keap1) was increased significantly, whereas heme oxygenase-1 (HO-1) was upregulated and NAD(P)H quinone oxidoreductase-1 (NQO1) was barely increased in the cytoplasm of tubules in kidneys after treatment with AAI. BARD significantly upregulated renal Nrf2, NQO1 and HO-1 expression and downregulated Keap1 expression compared with those in the AA group. Moreover, it was found that Nrf2 was expressed both in the cytoplasm and nuclear of glomeruli and tubules, whereas NQO1 and HO-1 were localized in the cytoplasm of tubules only. In conclusion, AA-induced acute renal injury was associated with impaired Nrf2 activation and expression of its downstream target genes in renal tissues. BARD prevented renal damage induced by AAI, and this renoprotective effect may be exerted by activating the Nrf2 signaling pathway and increasing expression of the downstream target genes. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Evolution of the biosynthesis of the branched-chain amino acids

Science.gov (United States)

Keefe, Anthony D.; Lazcano, Antonio; Miller, Stanley L.

1995-01-01

The origins of the biosynthetic pathways for the branched-chain amino acids cannot be understood in terms of the backwards development of the present acetolactate pathway because it contains unstable intermediates. We propose that the first biosynthesis of the branched-chain amino acids was by the reductive carboxylation of short branched chain fatty acids giving keto acids which were then transaminated. Similar reaction sequences mediated by nonspecific enzymes would produce serine and threomine from the abundant prebiotic compounds glycolic and lactic acids. The aromatic amino acids may also have first been synthesized in this way, e.g. tryptophan from indole acetic acid. The next step would have been the biosynthesis of leucine from alpha-ketoisovalerc acid. The acetolactate pathway developed subsequently. The first version of the Krebs cycle, which was used for amino acid biosynthesis, would have been assembled by making use fo the reductive carboxylation and leucine biosynthesis enzymes, and completed with the development of a single new enzyme, succinate dehydrogenase. This evolutionary scheme suggests that there may be limitations to inferring the origins of metabolism by a simple back extrapolation of current pathways.

10-Oxo-trans-11-octadecenoic acid generated from linoleic acid by a gut lactic acid bacterium Lactobacillus plantarum is cytoprotective against oxidative stress.

Science.gov (United States)

Furumoto, Hidehiro; Nanthirudjanar, Tharnath; Kume, Toshiaki; Izumi, Yasuhiko; Park, Si-Bum; Kitamura, Nahoko; Kishino, Shigenobu; Ogawa, Jun; Hirata, Takashi; Sugawara, Tatsuya

2016-04-01

Oxidative stress is a well-known cause of multiple diseases. The nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway plays a central role in cellular antioxidative responses. In this study, we investigated the effects of novel fatty acid metabolite derivatives of linoleic acid generated by the gut lactic acid bacteria Lactobacillus plantarum on the Nrf2-ARE pathway. 10-Oxo-trans-11-octadecenoic acid (KetoC) protected HepG2 cells from cytotoxicity induced by hydrogen peroxide. KetoC also significantly increased cellular Nrf2 protein levels, ARE-dependent transcription, and the gene expression of antioxidative enzymes such as heme oxygenase-1 (HO-1), glutamate-cysteine ligase modifier subunit (GCLM), and quinone oxidoreductase 1 (NQO1) in HepG2 cells. Additionally, a single oral dose administration of KetoC also increased antioxidative gene expression and protein levels of Nrf2 and HO-1 in mouse organs. Since other fatty acid metabolites and linoleic acid did not affect cellular antioxidative responses, the cytoprotective effect of KetoC may be because of its α,β-unsaturated carbonyl moiety. Collectively, our data suggested that KetoC activated the Nrf2-ARE pathway to enhance cellular antioxidative responses in vitro and in vivo, which further suggests that KetoC may prevent multiple diseases induced by oxidative stress. Copyright © 2016. Published by Elsevier Inc.

Trans-Fatty Acids Aggravate Obesity, Insulin Resistance and Hepatic Steatosis in C57BL/6 Mice, Possibly by Suppressing the IRS1 Dependent Pathway

Directory of Open Access Journals (Sweden)

Xiaona Zhao

2016-05-01

Full Text Available Trans-fatty acid consumption has been reported as a risk factor for metabolic disorders and targeted organ damages. Nonetheless, little is known about the roles and mechanisms of trans-fatty acids in obesity, insulin resistance (IR and hepatic steatosis. Adult C57BL/6 male mice were fed with four different diets for 20 weeks: normal diet (ND, high fat diet (HFD, low trans-fatty acids diet (LTD and high trans-fatty acid diet (HTD. The diet-induced metabolic disorders were assessed by evaluating body weight, glucose tolerance test, hepatic steatosis and plasma lipid profiles post 20-week diet. Histological (H&E, Oil-Red-O staining and western blot analysis were employed to assess liver steatosis and potential signaling pathways. After 20-weeks of diet, the body weights of the four groups were 29.61 ± 1.89 g (ND, 39.04 ± 4.27 g (HFD, 34.09 ± 2.62 g (LTD and 43.78 ± 4.27 g (HTD (p < 0.05, respectively. HFD intake significantly impaired glucose tolerance, which was impaired further in the mice consuming the HTD diet. The effect was further exacerbated by HTD diet. Moreover, the HTD group exhibited significantly more severe liver steatosis compared with HFD group possibly through regulating adipose triglyceride lipase. The group consuming the HTD also exhibited significantly reduced levels of IRS1, phosphor-PKC and phosphor-AKT. These results support our hypothesis that consumption of a diet high in trans-fatty acids induces higher rates of obesity, IR and hepatic steatosis in male C57BL/6 mice, possibly by suppressing the IRS1dependent pathway.

Complexity in Acid?Base Titrations: Multimer Formation Between Phosphoric Acids and Imines

OpenAIRE

Malm, Christian; Kim, Heejae; Wagner, Manfred; Hunger, Johannes

2017-01-01

Abstract Solutions of Br?nsted acids with bases in aprotic solvents are not only common model systems to study the fundamentals of proton transfer pathways but are also highly relevant to Br?nsted acid catalysis. Despite their importance the light nature of the proton makes characterization of acid?base aggregates challenging. Here, we track such acid?base interactions over a broad range of relative compositions between diphenyl phosphoric acid and the base quinaldine in dichloromethane, by u...

Observational Study of a French and Belgian Multicenter Cohort of 23 Patients Diagnosed in Adulthood With Mevalonate Kinase Deficiency.

Science.gov (United States)

Durel, Cécile-Audrey; Aouba, Achille; Bienvenu, Boris; Deshayes, Samuel; Coppéré, Brigitte; Gombert, Bruno; Acquaviva-Bourdain, Cécile; Hachulla, Eric; Lecomte, Frédéric; Touitou, Isabelle; Ninet, Jacques; Philit, Jean-Baptiste; Messer, Laurent; Brouillard, Marc; Girard-Madoux, Marie-Hélène; Moutschen, Michel; Raison-Peyron, Nadia; Hutin, Pascal; Duffau, Pierre; Trolliet, Pierre; Hatron, Pierre-Yves; Heudier, Philippe; Cevallos, Ramiro; Lequerré, Thierry; Brousse, Valentine; Lesire, Vincent; Audia, Sylvain; Maucort-Boulch, Delphine; Cuisset, Laurence; Hot, Arnaud

2016-03-01

The aim of this study was to describe the clinical and biological features of Mevalonate kinase deficiency (MKD) in patients diagnosed in adulthood. This is a French and Belgian observational retrospective study from 2000 to 2014. To constitute the cohort, we cross-check the genetic and biochemical databases. The clinical, enzymatic, and genetic data were gathered from medical records. Twenty-three patients were analyzed. The mean age at diagnosis was 40 years, with a mean age at onset of symptoms of 3 years. All symptomatic patients had fever. Febrile attacks were mostly associated with arthralgia (90.9%); lymphadenopathy, abdominal pain, and skin lesions (86.4%); pharyngitis (63.6%); cough (59.1%); diarrhea, and hepatosplenomegaly (50.0%). Seven patients had psychiatric symptoms (31.8%). One patient developed recurrent seizures. Three patients experienced renal involvement (13.6%). Two patients had angiomyolipoma (9.1%). All but one tested patients had elevated serum immunoglobulin (Ig) D level. Twenty-one patients had genetic diagnosis; most of them were compound heterozygote (76.2%). p.Val377Ile was the most prevalent mutation. Structural articular damages and systemic AA amyloidosis were the 2 most serious complications. More than 65% of patients displayed decrease in severity and frequency of attacks with increasing age, but only 35% achieved remission. MKD diagnosed in adulthood shared clinical and genetic features with classical pediatric disease. An elevated IgD concentration is a good marker for MKD in adults. Despite a decrease of severity and frequency of attacks with age, only one-third of patients achieved spontaneous remission.

Arachidonic Acid Metabolism Pathway Is Not Only Dominant in Metabolic Modulation but Associated With Phenotypic Variation After Acute Hypoxia Exposure

Directory of Open Access Journals (Sweden)

Chang Liu

2018-03-01

Full Text Available Background: The modulation of arachidonic acid (AA metabolism pathway is identified in metabolic alterations after hypoxia exposure, but its biological function is controversial. We aimed at integrating plasma metabolomic and transcriptomic approaches to systematically explore the roles of the AA metabolism pathway in response to acute hypoxia using an acute mountain sickness (AMS model.Methods: Blood samples were obtained from 53 enrolled subjects before and after exposure to high altitude. Ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry and RNA sequencing were separately performed for metabolomic and transcriptomic profiling, respectively. Influential modules comprising essential metabolites and genes were identified by weighted gene co-expression network analysis (WGCNA after integrating metabolic information with phenotypic and transcriptomic datasets, respectively.Results: Enrolled subjects exhibited diverse response manners to hypoxia. Combined with obviously altered heart rate, oxygen saturation, hemoglobin, and Lake Louise Score (LLS, metabolomic profiling detected that 36 metabolites were highly related to clinical features in hypoxia responses, out of which 27 were upregulated and nine were downregulated, and could be mapped to AA metabolism pathway significantly. Integrated analysis of metabolomic and transcriptomic data revealed that these dominant molecules showed remarkable association with genes in gas transport incapacitation and disorders of hemoglobin metabolism pathways, such as ALAS2, HEMGN. After detailed description of AA metabolism pathway, we found that the molecules of 15-d-PGJ2, PGA2, PGE2, 12-O-3-OH-LTB4, LTD4, LTE4 were significantly up-regulated after hypoxia stimuli, and increased in those with poor response manner to hypoxia particularly. Further analysis in another cohort showed that genes in AA metabolism pathway such as PTGES, PTGS1, GGT1, TBAS1 et al. were excessively

Branched-Chain Amino Acid Negatively Regulates KLF15 Expression via PI3K-AKT Pathway.

Science.gov (United States)

Liu, Yunxia; Dong, Weibing; Shao, Jing; Wang, Yibin; Zhou, Meiyi; Sun, Haipeng

2017-01-01

Recent studies have linked branched-chain amino acid (BCAA) with numerous metabolic diseases. However, the molecular basis of BCAA's roles in metabolic regulation remains to be established. KLF15 (Krüppel-like factor 15) is a transcription factor and master regulator of glycemic, lipid, and amino acids metabolism. In the present study, we found high concentrations of BCAA suppressed KLF15 expression while BCAA starvation induced KLF15 expression, suggesting KLF15 expression is negatively controlled by BCAA.Interestingly, BCAA starvation induced PI3K-AKT signaling. KLF15 induction by BCAA starvation was blocked by PI3K and AKT inhibitors, indicating the activation of PI3K-AKT signaling pathway mediated the KLF15 induction. BCAA regulated KLF15 expression at transcriptional level but not post-transcriptional level. However, BCAA starvation failed to increase the KLF15-promoter-driven luciferase expression, suggesting KLF15 promoter activity was not directly controlled by BCAA. Finally, fasting reduced BCAA abundance in mice and KLF15 expression was dramatically induced in muscle and white adipose tissue, but not in liver. Together, these data demonstrated BCAA negatively regulated KLF15 expression, suggesting a novel molecular mechanism underlying BCAA's multiple functions in metabolic regulation.

Branched-Chain Amino Acid Negatively Regulates KLF15 Expression via PI3K-AKT Pathway

Directory of Open Access Journals (Sweden)

Yunxia Liu

2017-10-01

Full Text Available Recent studies have linked branched-chain amino acid (BCAA with numerous metabolic diseases. However, the molecular basis of BCAA's roles in metabolic regulation remains to be established. KLF15 (Krüppel-like factor 15 is a transcription factor and master regulator of glycemic, lipid, and amino acids metabolism. In the present study, we found high concentrations of BCAA suppressed KLF15 expression while BCAA starvation induced KLF15 expression, suggesting KLF15 expression is negatively controlled by BCAA.Interestingly, BCAA starvation induced PI3K-AKT signaling. KLF15 induction by BCAA starvation was blocked by PI3K and AKT inhibitors, indicating the activation of PI3K-AKT signaling pathway mediated the KLF15 induction. BCAA regulated KLF15 expression at transcriptional level but not post-transcriptional level. However, BCAA starvation failed to increase the KLF15-promoter-driven luciferase expression, suggesting KLF15 promoter activity was not directly controlled by BCAA. Finally, fasting reduced BCAA abundance in mice and KLF15 expression was dramatically induced in muscle and white adipose tissue, but not in liver. Together, these data demonstrated BCAA negatively regulated KLF15 expression, suggesting a novel molecular mechanism underlying BCAA's multiple functions in metabolic regulation.

Unbiased plasma metabolomics reveal the correlation of metabolic pathways and Prakritis of humans.

Science.gov (United States)

Shirolkar, Amey; Chakraborty, Sutapa; Mandal, Tusharkanti; Dabur, Rajesh

2017-11-25

Ayurveda, an ancient Indian medicinal system, has categorized human body constitutions in three broad constitutional types (prakritis) i.e. Vata, Pitta and Kapha. Analysis of plasma metabolites and related pathways to classify Prakriti specific dominant marker metabolites and metabolic pathways. 38 healthy male individuals were assessed for dominant Prakritis and their fasting blood samples were collected. The processed plasma samples were subjected to rapid resolution liquid chromatography-electrospray ionization-quadrupole time of flight mass spectrometry (RRLC-ESI-QTOFMS). Mass profiles were aligned and subjected to multivariate analysis. Partial least square discriminant analysis (PLS-DA) model showed 97.87% recognition capability. List of PLS-DA metabolites was subjected to permutative Benjamini-Hochberg false discovery rate (FDR) correction and final list of 76 metabolites with p  2.0 was identified. Pathway analysis using metascape and JEPETTO plugins in Cytoscape revealed that steroidal hormone biosynthesis, amino acid, and arachidonic acid metabolism are major pathways varying with different constitution. Biological Go processes analysis showed that aromatic amino acids, sphingolipids, and pyrimidine nucleotides metabolic processes were dominant in kapha type of body constitution. Fat soluble vitamins, cellular amino acid, and androgen biosynthesis process along with branched chain amino acid and glycerolipid catabolic processes were dominant in pitta type individuals. Vata Prakriti was found to have dominant catecholamine, arachidonic acid and hydrogen peroxide metabolomics processes. The neurotransmission and oxidative stress in vata, BCAA catabolic, androgen, xenobiotics metabolic processes in pitta, and aromatic amino acids, sphingolipid, and pyrimidine metabolic process in kaphaPrakriti were the dominant marker pathways. Copyright © 2017 Transdisciplinary University, Bangalore and World Ayurveda Foundation. Published by Elsevier B.V. All rights

Dual pathways for ribonucleic acid turnover in WI-38 but not in I-cell human diploid fibroblasts

International Nuclear Information System (INIS)

Sameshima, M.; Liebhaber, S.A.; Schlessinger, D.

1981-01-01

The turnover rates of /sup 3/H-labeled 18S ribosomal ribonucleic acid (RNA), 28S ribsomal RNA, transfer RNA, and total cytoplasmic RNA were very similar in growing WI-38 diploid fibroblasts. The rate of turnover was at least twofold greater when cell growth stopped due to cell confluence, /sup 3/H irradiation, or treatment with 20 mM NaN/sub 3/ or 2 mM NaF. In contrast, the rate of total /sup 3/H-protein turnover was the same in growing and nongrowing cells. Both RNA and protein turnovers were accelerated at least twofold in WI-38 cells deprived of serum, and this increase in turnover was inhibited by NH/sub 4/Cl. These results are consistent with two pathways for RNA turnover, oe of them being nonlysosomal and the other being lyosome mediated (NH/sub 4/Cl sensitive), as has been suggested for protein turnover. Also consistent with the notion of two pathways for RNA turnover were findings with I-cells, which are deficient for many lysosomal enzymes, and in which all RNA turnover were nonlysosomal (NH/sub 4/Cl resistant)

Serum n-3 Tetracosapentaenoic Acid and Tetracosahexaenoic Acid Increase Following Higher Dietary α-Linolenic Acid but not Docosahexaenoic Acid.

Science.gov (United States)

Metherel, Adam H; Domenichiello, Anthony F; Kitson, Alex P; Lin, Yu-Hong; Bazinet, Richard P

2017-02-01

n-3 Tetracosapentaenoic acid (24:5n-3, TPAn-3) and tetracosahexaenoic acid (24:6n-3, THA) are believed to be important intermediates to docosahexaenoic acid (DHA, 22:6n-3) synthesis. The purpose of this study is to report for the first time serum concentrations of TPAn-3 and THA and their response to changing dietary α-linolenic acid (18:3n-3, ALA) and DHA. The responses will then be used in an attempt to predict the location of these fatty acids in relation to DHA in the biosynthetic pathway. Male Long Evans rats (n = 6 per group) were fed either a low (0.1% of total fatty acids), medium (3%) or high (10%) ALA diet with no added DHA, or a low (0%), medium (0.2%) or high (2%) DHA diet with a background of 2% ALA for 8 weeks post-weaning. Serum n-3 and n-6 polyunsaturated fatty acid (PUFA) concentrations (nmol/mL ± SEM) were determined by gas chromatography-mass spectrometry. Serum THA increases from low (0.3 ± 0.1) to medium (5.8 ± 0.7) but not from medium to high (4.6 ± 0.9) dietary ALA, while serum TPAn-3 increases with increasing dietary ALA from 0.09 ± 0.04 to 0.70 ± 0.09 to 1.23 ± 0.14 nmol/mL. Following DHA feeding, neither TPAn-3 or THA change across all dietary DHA intake levels. Serum TPAn-3 demonstrates a similar response to dietary DHA. In conclusion, this is the first study to demonstrate that increases in dietary ALA but not DHA increase serum TPAn-3 and THA in rats, suggesting that both fatty acids are precursors to DHA in the biosynthetic pathway.

Maternal smoking and the retinoid pathway in the developing lung

Directory of Open Access Journals (Sweden)

Manoli Sara E

2012-06-01

Full Text Available Abstract Background Maternal smoking is a risk factor for pediatric lung disease, including asthma. Animal models suggest that maternal smoking causes defective alveolarization in the offspring. Retinoic acid signaling modulates both lung development and postnatal immune function. Thus, abnormalities in this pathway could mediate maternal smoking effects. We tested whether maternal smoking disrupts retinoic acid pathway expression and functioning in a murine model. Methods Female C57Bl/6 mice with/without mainstream cigarette smoke exposure (3 research cigarettes a day, 5 days a week were mated to nonsmoking males. Cigarette smoke exposure continued throughout the pregnancy and after parturition. Lung tissue from the offspring was examined by mean linear intercept analysis and by quantitative PCR. Cell culture experiments using the type II cell-like cell line, A549, tested whether lipid-soluble cigarette smoke components affected binding and activation of retinoic acid response elements in vitro. Results Compared to tobacco-naïve mice, juvenile mice with tobacco toxin exposure had significantly (P  Conclusions A murine model of maternal cigarette smoking causes abnormal alveolarization in association with altered retinoic acid pathway element expression in the offspring. An in vitro cell culture model shows that lipid-soluble components of cigarette smoke decrease retinoic acid response element activation. It is feasible that disruption of retinoic acid signaling contributes to the pediatric lung dysfunction caused by maternal smoking.

Competitive fragmentation pathways of acetic acid dimer explored by synchrotron VUV photoionization mass spectrometry and electronic structure calculations.

Science.gov (United States)

Guan, Jiwen; Hu, Yongjun; Zou, Hao; Cao, Lanlan; Liu, Fuyi; Shan, Xiaobin; Sheng, Liusi

2012-09-28

In present study, photoionization and dissociation of acetic acid dimers have been studied with the synchrotron vacuum ultraviolet photoionization mass spectrometry and theoretical calculations. Besides the intense signal corresponding to protonated cluster ions (CH(3)COOH)(n)·H(+), the feature related to the fragment ions (CH(3)COOH)H(+)·COO (105 amu) via β-carbon-carbon bond cleavage is observed. By scanning photoionization efficiency spectra, appearance energies of the fragments (CH(3)COOH)·H(+) and (CH(3)COOH)H(+)·COO are obtained. With the aid of theoretical calculations, seven fragmentation channels of acetic acid dimer cations were discussed, where five cation isomers of acetic acid dimer are involved. While four of them are found to generate the protonated species, only one of them can dissociate into a C-C bond cleavage product (CH(3)COOH)H(+)·COO. After surmounting the methyl hydrogen-transfer barrier 10.84 ± 0.05 eV, the opening of dissociative channel to produce ions (CH(3)COOH)(+) becomes the most competitive path. When photon energy increases to 12.4 eV, we also found dimer cations can be fragmented and generate new cations (CH(3)COOH)·CH(3)CO(+). Kinetics, thermodynamics, and entropy factors for these competitive dissociation pathways are discussed. The present report provides a clear picture of the photoionization and dissociation processes of the acetic acid dimer in the range of the photon energy 9-15 eV.

Competitive fragmentation pathways of acetic acid dimer explored by synchrotron VUV photoionization mass spectrometry and electronic structure calculations

Science.gov (United States)

Guan, Jiwen; Hu, Yongjun; Zou, Hao; Cao, Lanlan; Liu, Fuyi; Shan, Xiaobin; Sheng, Liusi

2012-09-01

In present study, photoionization and dissociation of acetic acid dimers have been studied with the synchrotron vacuum ultraviolet photoionization mass spectrometry and theoretical calculations. Besides the intense signal corresponding to protonated cluster ions (CH3COOH)n.H+, the feature related to the fragment ions (CH3COOH)H+.COO (105 amu) via β-carbon-carbon bond cleavage is observed. By scanning photoionization efficiency spectra, appearance energies of the fragments (CH3COOH).H+ and (CH3COOH)H+.COO are obtained. With the aid of theoretical calculations, seven fragmentation channels of acetic acid dimer cations were discussed, where five cation isomers of acetic acid dimer are involved. While four of them are found to generate the protonated species, only one of them can dissociate into a C-C bond cleavage product (CH3COOH)H+.COO. After surmounting the methyl hydrogen-transfer barrier 10.84 ± 0.05 eV, the opening of dissociative channel to produce ions (CH3COOH)+ becomes the most competitive path. When photon energy increases to 12.4 eV, we also found dimer cations can be fragmented and generate new cations (CH3COOH).CH3CO+. Kinetics, thermodynamics, and entropy factors for these competitive dissociation pathways are discussed. The present report provides a clear picture of the photoionization and dissociation processes of the acetic acid dimer in the range of the photon energy 9-15 eV.

A Study on Enhanced Expression of 3-Hydroxypropionic Acid Pathway Genes and Impact on Its Production in Lactobacillus reuteri

Directory of Open Access Journals (Sweden)

Gopal Ramakrishnan Gopi

2015-01-01

Full Text Available 3-Hydroxypropionic acid (3-HP is a novel antimicrobial agent against foodborne pathogens like Salmonella and Staphylococcus species. Lactobacillus reuteri converts glycerol into 3-HP using a coenzyme A-dependent pathway, which is encoded by propanediol utilization operon (pdu subjected to catabolite repression. In a catabolite repression-deregulated L. reuteri RPRB3007, quantitative PCR revealed a 2.5-fold increase in the transcripts of the genes pduP, pduW and pduL during the mid-log phase of growth. The production of 3-HP was tested in resting cells in phosphate buff er and growing batch cultures in MRS broth of various glucose/glycerol ratios. Due to the upregulation of pathway genes, specific formation rate of 3-HP in the mutant strain was found to be enhanced from 0.167 to 0.257 g per g of cell dry mass per h. Furthermore, formation of 3-HP in resting cells was limited due to the substrate inhibition by reuterin at a concentration of (30±5 mM. In batch cultures, the formation of 3-HP was not observed during the logarithmic and stationary phases of growth of wild-type and mutant strains, which was confi rmed by NMR spectroscopy. However, the cells collected in these phases were found to produce 3-HP aft er washing and converting them to resting cells. Lactate and acetate, the primary end products of glucose catabolism, might be the inhibiting elements for 3-HP formation in batch cultures. This was confirmed when lactate (25±5 mM or acetate (20±5 mM were added to biotransformation medium, which prevented the 3-HP formation. Moreover, the removal of sodium acetate and glucose (carbon source for lactic acid production was found to restore 3-HP formation in the MRS broth in a similar manner to that of the phosphate buff er. Even though the genetic repression was circumvented by the up-regulation of pathway genes using a mutant strain, 3-HP formation was further limited by the substrate and catabolite inhibition.

Chemical and physical structures of proteinoids and related polyamino acids

Science.gov (United States)

Mita, Hajime; Kuwahara, Yusuke; Nomoto, Shinya

Studies of polyamino acid formation pathways in the prebiotic condition are important for the study of the origins of life. Several pathways of prebiotic polyamino acid formation have been reported. Heating of monoammonium malate [1] and heating of amino acids in molten urea [2] are important pathways of the prebiotic peptide formation. The former case, globular structure called proteinoid microsphere is formed in aqueous conditions. The later case, polyamino acids are formed from unrestricted amino acid species. Heating of aqueous aspargine is also interesting pathway for the prebiotic polyamino acid formation, because polyamino acid formation proceeds in aqueous condition [3]. In this study, we analyzed the chemical structure of the proteinoids and related polyamino acids formed in the above three pathways using with mass spectrometer. In addition, their physical structures are analyzed by the electron and optical microscopes, in order to determine the self-organization abilities. We discuss the relation between the chemical and the physical structures for the origins of life. References [1] Harada, K., J. Org. Chem., 24, 1662 (1959), Fox, S. W., Harada, K., and Kendrick, J., Science, 129, 1221 (1959). [2] Terasaki, M., Nomoto, S., Mita, H., and Shimoyama, A., Chem. Lett., 480 (2002), Mita, H., Nomoto, S., Terasaki, M., Shimoyama, A., and Yamamoto, Y., Int. J. Astrobiol., 4, 145 (2005). [3] Kovacs, K and Nagy, H., Nature, 190, 531 (1961), Munegumi, T., Tanikawa, N., Mita, H. and Harada, K., Viva Origino, 22, 109 (1994).

Insulin-dependent signaling: regulation by amino acids and energy

NARCIS (Netherlands)

Meijer, A. J.

2004-01-01

Recent research has indicated that amino acids stimulate a signal-transduction pathway that is also used by insulin. Moreover, for insulin to exert its anabolic and anticatabolic effects on protein, there is an absolute requirement for amino acids. This signaling pathway becomes inhibited by

Ectopic expression of Crambe abyssinica lysophosphatidic acid ...

African Journals Online (AJOL)

USER

2010-06-21

Jun 21, 2010 ... lysophosphatidic acid acyltransferase in transgenic rapeseed increases its oil .... pathway [fatty acid desaturase-2 (BnFAD2, AY577313), fatty acid desaturase-3 ..... Acyltransferases from basic science to modified seed oils.

In vitro reconstitution of an abscisic acid signalling pathway

KAUST Repository

Fujii, Hiroaki; Chinnusamy, Viswanathan; Rodrigues, Americo; Rubio, Silvia; Antoni, Regina; Park, Sang-Youl; Cutler, Sean R.; Sheen, Jen; Rodriguez, Pedro L.; Zhu, Jian-Kang

2009-01-01

The phytohormone abscisic acid (ABA) regulates the expression of many genes in plants; it has critical functions in stress resistance and in growth and development. Several proteins have been reported to function as ABA receptors, and many more are known to be involved in ABA signalling. However, the identities of ABA receptors remain controversial and the mechanism of signalling from perception to downstream gene expression is unclear. Here we show that by combining the recently identified ABA receptor PYR1 with the type 2C protein phosphatase (PP2C) ABI1, the serine/threonine protein kinase SnRK2.6/OST1 and the transcription factor ABF2/AREB1, we can reconstitute ABA-triggered phosphorylation of the transcription factor in vitro. Introduction of these four components into plant protoplasts results in ABA-responsive gene expression. Protoplast and test-tube reconstitution assays were used to test the function of various members of the receptor, protein phosphatase and kinase families. Our results suggest that the default state of the SnRK2 kinases is an autophosphorylated, active state and that the SnRK2 kinases are kept inactive by the PP2Cs through physical interaction and dephosphorylation. We found that in the presence of ABA, the PYR/PYL (pyrabactin resistance 1/PYR1-like) receptor proteins can disrupt the interaction between the SnRK2s and PP2Cs, thus preventing the PP2C-mediated dephosphorylation of the SnRK2s and resulting in the activation of the SnRK2 kinases. Our results reveal new insights into ABA signalling mechanisms and define a minimal set of core components of a complete major ABA signalling pathway. © 2009 Macmillan Publishers Limited. All rights reserved.

In vitro reconstitution of an abscisic acid signalling pathway

KAUST Repository

Fujii, Hiroaki

2009-11-18

The phytohormone abscisic acid (ABA) regulates the expression of many genes in plants; it has critical functions in stress resistance and in growth and development. Several proteins have been reported to function as ABA receptors, and many more are known to be involved in ABA signalling. However, the identities of ABA receptors remain controversial and the mechanism of signalling from perception to downstream gene expression is unclear. Here we show that by combining the recently identified ABA receptor PYR1 with the type 2C protein phosphatase (PP2C) ABI1, the serine/threonine protein kinase SnRK2.6/OST1 and the transcription factor ABF2/AREB1, we can reconstitute ABA-triggered phosphorylation of the transcription factor in vitro. Introduction of these four components into plant protoplasts results in ABA-responsive gene expression. Protoplast and test-tube reconstitution assays were used to test the function of various members of the receptor, protein phosphatase and kinase families. Our results suggest that the default state of the SnRK2 kinases is an autophosphorylated, active state and that the SnRK2 kinases are kept inactive by the PP2Cs through physical interaction and dephosphorylation. We found that in the presence of ABA, the PYR/PYL (pyrabactin resistance 1/PYR1-like) receptor proteins can disrupt the interaction between the SnRK2s and PP2Cs, thus preventing the PP2C-mediated dephosphorylation of the SnRK2s and resulting in the activation of the SnRK2 kinases. Our results reveal new insights into ABA signalling mechanisms and define a minimal set of core components of a complete major ABA signalling pathway. © 2009 Macmillan Publishers Limited. All rights reserved.

The application of glutamic acid alpha-decarboxylase for the valorization of glutamic acid

NARCIS (Netherlands)

Lammens, T.M.; Biase, De Daniela; Franssen, M.C.R.; Scott, E.L.; Sanders, J.P.M.

2009-01-01

Glutamic acid is an important constituent of waste streams from biofuels production. It is an interesting starting material for the synthesis of nitrogen containing bulk chemicals, thereby decreasing the dependency on fossil fuels. On the pathway from glutamic acid to a range of molecules, the

Low-ω3 Fatty Acid and Soy Protein Attenuate Alcohol-Induced Fatty Liver and Injury by Regulating the Opposing Lipid Oxidation and Lipogenic Signaling Pathways

Directory of Open Access Journals (Sweden)

Karina Reyes-Gordillo

2016-01-01

Full Text Available Chronic ethanol-induced downregulation of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α and upregulation of peroxisome proliferator-activated receptor gamma coactivator 1-beta (PGC1β affect hepatic lipid oxidation and lipogenesis, respectively, leading to fatty liver injury. Low-ω3 fatty acid (Low-ω3FA that primarily regulates PGC1α and soy protein (SP that seems to have its major regulatory effect on PGC1β were evaluated for their protective effects against ethanol-induced hepatosteatosis in rats fed with Lieber-deCarli control or ethanol liquid diets with high or low ω3FA fish oil and soy protein. Low-ω3FA and SP opposed the actions of chronic ethanol by reducing serum and liver lipids with concomitant decreased fatty liver. They also prevented the downregulation of hepatic Sirtuin 1 (SIRT1 and PGC1α and their target fatty acid oxidation pathway genes and attenuated the upregulation of hepatic PGC1β and sterol regulatory element-binding protein 1c (SREBP1c and their target lipogenic pathway genes via the phosphorylation of 5′ adenosine monophosphate-activated protein kinase (AMPK. Thus, these two novel modulators attenuate ethanol-induced hepatosteatosis and consequent liver injury potentially by regulating the two opposing lipid oxidation and lipogenic pathways.

Selective deoxygenation of stearic acid via an anhydride pathway

NARCIS (Netherlands)

Hollak, S.A.W.; Bitter, W.; Haveren, van J.; Es, van D.S.

2012-01-01

Stearic anhydride is proposed as reactive intermediate in the hydrogen free decarbonylation and ketonization of stearic acid over Pd/Al2O3 at 523 K. This information is crucial towards developing of a selective low temperature decarbonylation process of fatty acids towards olefins.

Enzymatic synthesis of 11C-pyruvic acid and 11C-L-lactic acid

International Nuclear Information System (INIS)

Cohen, M.B.; Spolter, L.; Chang, C.C.; Cook, J.S.; Macdonald, N.S.

1980-01-01

L-Lactic acid is formed as the end product of glycolysis under anaerobic conditions in all cells, but this reaction is of special significance in the myocardium. L-Lactic acid is reversibly formed from and is in equilibrium with myocardial pyruvic acid, which is its sole metabolic pathway. 11 C-Pyruvic acid is synthesized from 11 C carbon dioxide using pyruvate-ferredoxin oxidoreductase and coenzymes. The 11 C-pyruvic acid is then converted to 11 -L-lactic acid by lactic acid dehydrogenase. The availability of 11 C-pyruvic acid and 11 C-L-lactic acid will permit the in vivo investigation of lactate metabolism. (author)

Imbalanced Kynurenine Pathway in Schizophrenia

Directory of Open Access Journals (Sweden)

Magdalena E. Kegel

2014-01-01

Full Text Available Several studies suggest a role for kynurenic acid (KYNA in the pathophysiology of schizophrenia. It has been proposed that increased brain KYNA levels in schizophrenia result from a pathological shift in the kynurenine pathway toward enhanced KYNA formation, away from the other branch of the pathway leading to quinolinic acid (QUIN. Here we investigate the levels of QUIN in cerebrospinal fluid (CSF of patients with schizophrenia and healthy controls, and relate those to CSF levels of KYNA and other kynurenine metabolites from the same individuals. CSF QUIN levels from stable outpatients treated with olanzapine (n = 22 and those of controls (n = 26 were analyzed using liquid chromatography-mass spectrometry. No difference in CSF QUIN levels between patients and controls was observed (20.6 ± 1.5 nM vs. 18.2 ± 1.1 nM, P = 0.36. CSF QUIN was positively correlated to CSF kynurenine and CSF KYNA in patients but not in controls. The CSF QUIN/KYNA ratio was lower in patients than in controls ( P = 0.027. In summary, the present study offers support for an over-activated and imbalanced kynurenine pathway, favoring the production of KYNA over QUIN in patients with schizophrenia.

Dietary influence on MAPK-signaling pathways and risk of colon and rectal cancer.

Science.gov (United States)

Slattery, Martha L; Lundgreen, Abbie; Wolff, Roger K

2013-01-01

Mitogen-activated protein kinase (MAPK) pathways regulate cellular functions including cell proliferation, differentiation, migration, and apoptosis. Associations between genes in the DUSP, ERK1/2, JNK, and p38 MAPK-signaling pathways and dietary factors associated with growth factors, inflammation, and oxidative stress and risk of colon and rectal cancer were evaluated. Data include colon cases (n = 1555) and controls (n = 1956) and rectal cases (n = 754) and controls (n = 959). Statistically significant interactions were observed for the MAPK-signaling pathways after adjustment for multiple comparisons. DUSP genes interacted with carbohydrates, mutagen index, calories, calcium, vitamin D, lycopene, dietary fats, folic acid, and selenium. MAPK1, MAPK3, MAPK1, and RAF1 within the ERK1/2 MAPK-signaling pathway interacted with dietary fats and cruciferous vegetables. Within the JNK MAPK-signaling pathway, interactions between MAP3K7 and protein, vitamin C, iron, folic acid, carbohydrates, and cruciferous vegetables; MAP3K10 and folic acid; MAP3K9 and lutein/zeaxanthin; MAPK8 and calcium; MAP3K3 and calcium and lutein; MAP3K1 and cruciferous vegetables. Interaction within the p38-signaling pathway included MAPK14 with calories, carbohydrates saturated fat, selenium, vitamin C; MAP3K2 and carbohydrates, and folic acid. These data suggest that dietary factors involved in inflammation and oxidative stress interact with MAPK-signaling genes to alter risk of colorectal cancer.

Nucleotide Metabolism and its Control in Lactic Acid Bacteria

DEFF Research Database (Denmark)

Kilstrup, Mogens; Hammer, Karin; Jensen, Peter Ruhdal

2005-01-01

Most metabolic reactions are connected through either their utilization of nucleotides or their utilization of nucleotides or their regulation by these metabolites. In this review the biosynthetic pathways for pyrimidine and purine metabolism in lactic acid bacteria are described including...... the interconversion pathways, the formation of deoxyribonucleotides and the salvage pathways for use of exogenous precursors. The data for the enzymatic and the genetic regulation of these pathways are reviewed, as well as the gene organizations in different lactic acid bacteria. Mutant phenotypes and methods...... for manipulation of nucleotide pools are also discussed. Our aim is to provide an overview of the physiology and genetics of nucleotide metabolism and its regulation that will facilitate the interpretation of data arising from genetics, metabolomics, proteomics, and transcriptomics in lactic acid bacteria....

Zurampic Protects Pancreatic β-Cells from High Uric Acid Induced-Damage by Inhibiting URAT1 and Inactivating the ROS/AMPK/ERK Pathways

Directory of Open Access Journals (Sweden)

Ying Xin

2018-05-01

Full Text Available Background/Aims: Zurampic is a US FDA approved drug for treatment of gout. However, the influence of Zurampic on pancreatic β-cells remains unclear. The study aimed to evaluate the effects of Zurampic on high uric acid-induced damage of pancreatic β-cells and the possible underlying mechanisms. Methods: INS-1 cells and primary rat islets were stimulated with Zurampic and the mRNA expression of urate transporter 1 (URAT1 was assessed by qRT-PCR. Cells were stimulated with uric acid or uric acid plus Zurampic, and cell viability, apoptosis and ROS release were measured by MTT and flow cytometry assays. Western blot analysis was performed to evaluate the expressions of active Caspase-3 and phosphorylation of AMPK and ERK. Finally, cells were stimulated with uric acid or uric acid plus Zurampic at low/high level of glucose (2.8/16.7 mM glucose, and the insulin release was assessed by ELISA. Results: mRNA expression of URAT1 was decreased by Zurampic in a dose-dependent manner. Uric acid decreased cell viability, promoted cell apoptosis and induced ROS release. Uric acid-induced alterations could be reversed by Zurampic. Activation of Caspase-3 and phosphorylation of AMPK and ERK were enhanced by uric acid, and the enhancements were reversed by Zurampic. Decreased phosphorylation of AMPK and ERK, induced by Zurampic, was further reduced by adding inhibitor of AMPK or ERK. Besides, uric acid inhibited high glucose-induced insulin secretion and the inhibition was rescued by Zurampic. Conclusions: Zurampic has a protective effect on pancreatic β-cells against uric acid induced-damage by inhibiting URAT1 and inactivating the ROS/AMPK/ERK pathway.

Olfactory transduction pathways in the Senegalese sole Solea senegalensis.

Science.gov (United States)

Velez, Z; Hubbard, P C; Barata, E N; Canário, A V M

2013-09-01

This study tested whether differences in sensitivity between the upper and lower olfactory epithelia of Solea senegalensis are associated with different odorant receptors and transduction pathways, using the electro-olfactogram. Receptor mechanisms were assessed by cross-adaptation with amino acids (L-cysteine, L-phenylalanine and 1-methyl-L-tryptophan) and bile acids (taurocholic acid and cholic acid). This suggested that relatively specific receptors exist for 1-methyl-L-tryptophan and L-phenylalanine (food-related odorants) in the lower epithelium, and for taurocholic acid (conspecific-derived odorant) in the upper. Inhibition by U73122 [a phospholipase C (PLC) inhibitor] suggested that olfactory responses to amino acids were mediated mostly, but not entirely, by PLC-mediated transduction (IC50 ; 15-55 nM), whereas bile acid responses were mediated by both PLC and adenylate cyclase-cyclic adenosine monophosphate (AC-cAMP) (using SQ-22536; an AC inhibitor). Simultaneous application of both drugs rarely inhibited responses completely, suggesting possible involvement of non-PLC and non-AC mediated mechanisms. For aromatic amino acids and bile acids, there were differences in the contribution of each transduction pathway (PLC, AC and non-PLC and non-AC) between the two epithelia. These results suggest that differences in sensitivity of the two epithelia are associated with differences in odorant receptors and transduction mechanisms. © 2013 The Fisheries Society of the British Isles.

Effects of Peptone Supplementation in Different Culture Media on Growth, Metabolic Pathway and Productivity of CHO DG44 Cells; a New Insight into Amino Acid Profiles.

Science.gov (United States)

Davami, Fatemeh; Eghbalpour, Farnaz; Nematollahi, Leila; Barkhordari, Farzaneh; Mahboudi, Fereidoun

2015-01-01

The optimization of bioprocess conditions towards improved growth profile and productivity yield is considered of great importance in biopharmaceutical manufacturing. Peptones as efficient sources of nutrients have been studied for their effect on media development; however, their role on metabolic pathway is not well understood. In the present study, the effect of different concentration of peptones on a recombinant Chinese hamster ovary (CHO) cell line grown in three serum-free suspension cultures was determined. Six peptones of different origins and available amino acid profiles were investigated regarding their impact on cell growth, productivity, and metabolic pathways changes. In optimized feeding strategies, increases of 136% and 159% in volumetric productivity (for a low-nutrient culture media) and 55% (for a high-nutrient culture media) were achieved. Furthermore, particular sources of peptones with specific amino acid profile developed preferential results for each different culture medium. Two peptones, SoyA2SC and SoyE-110, were the only hydrolysates that showed production improvement in all three media. Casein Peptone plus Tryptone N1 and SoyA3SC showed different improved results based on their implemented concentration for each individual basal medium. The amino acid profile of peptones may provide clues to identify the most effective feeding strategies for recombinant CHO cells.

Pathway Analysis of miR-181a in Hypopharyngeal Squamous Cell ...

African Journals Online (AJOL)

syakima

2012-03-15

Mar 15, 2012 ... Key words: MicroRNA, head and neck cancer, miR-181a, pathway analysis, luciferase assay, FaDu cell line, transfection ..... prostate carcinoma and glioblastoma cells. Nucleic Acids .... WNT pathway in oral cancer: epigenetic.

Requirement of a Functional Flavin Mononucleotide Prenyltransferase for the Activity of a Bacterial Decarboxylase in a Heterologous Muconic Acid Pathway in Saccharomyces cerevisiae.

Science.gov (United States)

Weber, Heike E; Gottardi, Manuela; Brückner, Christine; Oreb, Mislav; Boles, Eckhard; Tripp, Joanna

2017-05-15

Biotechnological production of cis , cis -muconic acid from renewable feedstocks is an environmentally sustainable alternative to conventional, petroleum-based methods. Even though a heterologous production pathway for cis , cis -muconic acid has already been established in the host organism Saccharomyces cerevisiae , the generation of industrially relevant amounts of cis , cis -muconic acid is hampered by the low activity of the bacterial protocatechuic acid (PCA) decarboxylase AroY isomeric subunit C iso (AroY-C iso ), leading to secretion of large amounts of the intermediate PCA into the medium. In the present study, we show that the activity of AroY-C iso in S. cerevisiae strongly depends on the strain background. We could demonstrate that the strain dependency is caused by the presence or absence of an intact genomic copy of PAD1 , which encodes a mitochondrial enzyme responsible for the biosynthesis of a prenylated form of the cofactor flavin mononucleotide (prFMN). The inactivity of AroY-C iso in strain CEN.PK2-1 could be overcome by plasmid-borne expression of Pad1 or its bacterial homologue AroY subunit B (AroY-B). Our data reveal that the two enzymes perform the same function in decarboxylation of PCA by AroY-C iso , although coexpression of Pad1 led to higher decarboxylase activity. Conversely, AroY-B can replace Pad1 in its function in decarboxylation of phenylacrylic acids by ferulic acid decarboxylase Fdc1. Targeting of the majority of AroY-B to mitochondria by fusion to a heterologous mitochondrial targeting signal did not improve decarboxylase activity of AroY-C iso , suggesting that mitochondrial localization has no major impact on cofactor biosynthesis. IMPORTANCE In Saccharomyces cerevisiae , the decarboxylation of protocatechuic acid (PCA) to catechol is the bottleneck reaction in the heterologous biosynthetic pathway for production of cis , cis -muconic acid, a valuable precursor for the production of bulk chemicals. In our work, we demonstrate

Lactobacillus bulgaricus OLL1181 activates the aryl hydrocarbon receptor pathway and inhibits colitis

Science.gov (United States)

Takamura, Takeyuki; Harama, Daisuke; Fukumoto, Suguru; Nakamura, Yuki; Shimokawa, Naomi; Ishimaru, Kayoko; Ikegami, Shuji; Makino, Seiya; Kitamura, Masanori; Nakao, Atsuhito

2011-01-01

Increasing evidence suggests that the aryl hydrocarbon receptor (AhR) pathway has an important role in the regulation of inflammatory responses. Most recently, we have shown that the activation of the AhR pathway by a potent AhR agonist inhibits the development of dextran sodium sulfate (DSS)-induced colitis, a model of human ulcerative colitis, by the induction of prostaglandin E2 (PGE2) in the large intestine. Because several strains of probiotic lactic acid bacteria have been reported to inhibit DSS-induced colitis by unidentified mechanisms, we hypothesized that particular strains of lactic acid bacterium might have the potential to activate the AhR pathway, thereby inhibiting DSS-induced colitis. This study investigated whether there are specific lactic acid bacterial strains that can activate the AhR pathway, and if so, whether this AhR-activating potential is associated with suppression of DSS-induced colitis. By using AhR signaling reporter cells, we found that Lactobacillus bulgaricus OLL1181 had the potential to activate the AhR pathway. OLL1181 also induced the mRNA expression of cytochrome P450 family 1A1 (CYP1A1), a target gene of the AhR pathway, in human colon cells, which was inhibited by the addition of an AhR antagonist, α-naphthoflavon (αNF). In addition, mice treated orally with OLL1181 showed an increase in CYP1A1 mRNA expression in the large intestine and amelioration of DSS-induced colitis. Thus, OLL1181 can induce activation of the intestinal AhR pathway and inhibit DSS-induced colitis in mice. This strain of lactic acid bacterium has therefore the potential to activate the AhR pathway, which may be able to suppress colitis. PMID:21321579

Streptozotocin induced activation of oxidative stress responsive splenic cell signaling pathways: Protective role of arjunolic acid

International Nuclear Information System (INIS)

Manna, Prasenjit; Ghosh, Jyotirmoy; Das, Joydeep; Sil, Parames C.

2010-01-01

Present study investigates the beneficial role of arjunolic acid (AA) against the alteration in the cytokine levels and simultaneous activation of oxidative stress responsive signaling pathways in spleen under hyperglycemic condition. Diabetes was induced by injection of streptozotocin (STZ) (at a dose of 70 mg/kg body weight, injected in the tail vain). STZ administration elevated the levels of IL-2 as well as IFN-γ and attenuated the level of TNF-α in the sera of diabetic animals. In addition, hyperglycemia is also associated with the increased production of intracellular reactive intermediates resulting with the elevation in lipid peroxidation, protein carbonylation and reduction in intracellular antioxidant defense. Investigating the oxidative stress responsive cell signaling pathways, increased expressions (immunoreactive concentrations) of phosphorylated p65 as well as its inhibitor protein phospho IκBα and phosphorylated mitogen activated protein kinases (MAPKs) have been observed in diabetic spleen tissue. Studies on isolated splenocytes revealed that hyperglycemia caused disruption of mitochondrial membrane potential, elevation in the concentration of cytosolic cytochrome c as well as activation of caspase 3 leading to apoptotic cell death. Histological examination revealed that diabetic induction depleted the white pulp scoring which is in agreement with the reduced immunological response. Treatment with AA prevented the hyperglycemia and its associated pathogenesis in spleen tissue. Results suggest that AA might act as an anti-diabetic and immunomodulatory agent against hyperglycemia.

Formation of biologically relevant carboxylic acids during the gamma irradiation of acetic acid

Science.gov (United States)

Negron-Mendoza, A.; Ponnamperuma, C.

1976-01-01

Irradiation of aqueous solutions of acetic acid with gamma rays produced several carboxylic acids in small yield. Their identification was based on the technique of gas chromatography combined with mass spectrometry. Some of these acids are Krebs Cycle intermediates. Their simultaneous formation in experiments simulating the primitive conditions on the earth suggests that metabolic pathways may have had their origin in prebiotic chemical processes.

Induction of Shikimic Acid Pathway Enzymes by Light in Suspension Cultured Cells of Parsley (Petroselinum crispum) 1

Science.gov (United States)

McCue, Kent F.; Conn, Eric E.

1990-01-01

Light treatment of suspension cultured cells of parsley (Petroselinum crispum) was shown to increase the activity of the shikimic acid pathway enzyme, 3-deoxy-d-arabino-heptulosonic acid-7-phosphate (DAHP) synthase (EC 4.1.2.15). DAHP synthase activity was assayed for two isoforms, DS-Mn and DS-Co (RJ Ganson, TA d'Amato, RA Jensen [1986] Plant Physiol 82: 203-210). Light increased the enzymatic activity of the plastidic isoform DS-Mn as much as 2-fold, averaging 1.6-fold with >95% confidence. The cytosolic isoform DS-Co was unaffected. Cycloheximide and actinomycin D, translational and transcriptional inhibitors, respectively, both reversed induction of DS-Mn by light suggesting transcriptional regulation of the gene. Chorismate mutase activity was assayed for the two isoforms CM I and CM II (BK Singh, JA Connelly, EE Conn [1985] Arch Biochem Biophys 243: 374-384). Treatment by light did not significantly affect either chorismate mutase isoform. The ratio of the two chorismate mutase isoforms changed during the growth cycle, with an increase in the ratio of plastidic to cytosolic isoforms occurring towards the end of logarithmic growth. PMID:16667741

FGF19 regulates cell proliferation, glucose and bile acid metabolism via FGFR4-dependent and independent pathways.

Directory of Open Access Journals (Sweden)

Ai-Luen Wu

Full Text Available Fibroblast growth factor 19 (FGF19 is a hormone-like protein that regulates carbohydrate, lipid and bile acid metabolism. At supra-physiological doses, FGF19 also increases hepatocyte proliferation and induces hepatocellular carcinogenesis in mice. Much of FGF19 activity is attributed to the activation of the liver enriched FGF Receptor 4 (FGFR4, although FGF19 can activate other FGFRs in vitro in the presence of the coreceptor βKlotho (KLB. In this report, we investigate the role of FGFR4 in mediating FGF19 activity by using Fgfr4 deficient mice as well as a variant of FGF19 protein (FGF19v which is specifically impaired in activating FGFR4. Our results demonstrate that FGFR4 activation mediates the induction of hepatocyte proliferation and the suppression of bile acid biosynthesis by FGF19, but is not essential for FGF19 to improve glucose and lipid metabolism in high fat diet fed mice as well as in leptin-deficient ob/ob mice. Thus, FGF19 acts through multiple receptor pathways to elicit pleiotropic effects in regulating nutrient metabolism and cell proliferation.

Mechanistic Perspectives of Maslinic Acid in Targeting Inflammation

Directory of Open Access Journals (Sweden)

Wei Hsum Yap

2015-01-01

Full Text Available Chronic inflammation drives the development of various pathological diseases such as rheumatoid arthritis, atherosclerosis, multiple sclerosis, and cancer. The arachidonic acid pathway represents one of the major mechanisms for inflammation. Prostaglandins (PGs are lipid products generated from arachidonic acid by the action of cyclooxygenase (COX enzymes and their activity is blocked by nonsteroidal anti-inflammatory drugs (NSAIDS. The use of natural compounds in regulation of COX activity/prostaglandins production is receiving increasing attention. In Mediterranean diet, olive oil and table olives contain significant dietary sources of maslinic acid. Maslinic acid is arising as a safe and novel natural pentacyclic triterpene which has protective effects against chronic inflammatory diseases in various in vivo and in vitro experimental models. Understanding the anti-inflammatory mechanism of maslinic acid is crucial for its development as a potential dietary nutraceutical. This review focuses on the mechanistic action of maslinic acid in regulating the inflammation pathways through modulation of the arachidonic acid metabolism including the nuclear factor-kappa B (NF-κB/COX-2 expression, upstream protein kinase signaling, and phospholipase A2 enzyme activity. Further investigations may provide insight into the mechanism of maslinic acid in regulating the molecular targets and their associated pathways in response to specific inflammatory stimuli.

Curcumin improves alcoholic fatty liver by inhibiting fatty acid biosynthesis.

Science.gov (United States)

Guo, Chang; Ma, Jingfan; Zhong, Qionghong; Zhao, Mengyuan; Hu, Tianxing; Chen, Tong; Qiu, Longxin; Wen, Longping

2017-08-01

Alcoholic fatty liver is a threat to human health. It has been long known that abstinence from alcohol is the most effective therapy, other effective therapies are not available for the treatment in humans. Curcumin has a great potential for anti-oxidation and anti-inflammation, but the effect on metabolic reconstruction remains little known. Here we performed metabolomic analysis by gas chromatography/mass spectrometry and explored ethanol pathogenic insight as well as curcumin action pattern. We identified seventy-one metabolites in mouse liver. Carbohydrates and lipids were characteristic categories. Pathway analysis results revealed that ethanol-induced pathways including biosynthesis of unsaturated fatty acids, fatty acid biosynthesis and pentose and glucuronate interconversions were suppressed by curcumin. Additionally, ethanol enhanced galactose metabolism and pentose phosphate pathway. Glyoxylate and dicarboxylate metabolism and pyruvate metabolism were inhibited in mice fed ethanol diet plus curcumin. Stearic acid, oleic acid and linoleic acid were disease biomarkers and therapical biomarkers. These results reflect the landscape of hepatic metabolism regulation. Our findings illustrate ethanol pathological pathway and metabolic mechanism of curcumin therapy. Copyright © 2017. Published by Elsevier Inc.

UV-C-Induced alleviation of transcriptional gene silencing through plant–plant communication: Key roles of jasmonic acid and salicylic acid pathways

Energy Technology Data Exchange (ETDEWEB)

Xu, Wei; Wang, Ting [Key Laboratory of Ion Beam Bio-engineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, P.O. Box 1138, Hefei, Anhui, 230031 (China); Xu, Shaoxin [School of physics and materials science, Anhui University, Hefei, Anhui, 230601 (China); Li, Fanghua; Deng, Chenguang; Wu, Lijun; Wu, Yuejin [Key Laboratory of Ion Beam Bio-engineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, P.O. Box 1138, Hefei, Anhui, 230031 (China); Bian, Po, E-mail: bianpo@ipp.ac.cn [Key Laboratory of Ion Beam Bio-engineering, Hefei Institutes of Physical Science, Chinese Academy of Sciences, P.O. Box 1138, Hefei, Anhui, 230031 (China)

2016-08-15

Highlights: • Transcriptional gene silencing (TGS) in plants can be epigenetically alleviated by volatile signals from UV-C- irradiated neighboring plants. • Alleviation of TGS can be induced by UV-C irradiation through plant–plant–plant communication. • JA and SA signals take part in interplant communication for alleviation of TGS. - Abstract: Plant stress responses at the epigenetic level are expected to allow more permanent changes of gene expression and potentially long-term adaptation. While it has been reported that plants subjected to adverse environments initiate various stress responses in their neighboring plants, little is known regarding epigenetic responses to external stresses mediated by plant–plant communication. In this study, we show that DNA repetitive elements of Arabidopsis thaliana, whose expression is inhibited epigenetically by transcriptional gene silencing (TGS) mechanism, are activated by UV-C irradiation through airborne plant–plant and plant–plant–plant communications, accompanied by DNA demethylation at CHH sites. Moreover, the TGS is alleviated by direct treatments with exogenous methyl jasmonate (MeJA) and methyl salicylate (MeSA). Further, the plant–plant and plant–plant–plant communications are blocked by mutations in the biosynthesis or signaling of jasmonic acid (JA) or salicylic acid (SA), indicating that JA and SA pathways are involved in the interplant communication for epigenetic responses. For the plant–plant–plant communication, stress cues are relayed to the last set of receiver plants by promoting the production of JA and SA signals in relaying plants, which exhibit upregulated expression of genes for JA and SA biosynthesis and enhanced emanation of MeJA and MeSA.

UV-C-Induced alleviation of transcriptional gene silencing through plant–plant communication: Key roles of jasmonic acid and salicylic acid pathways

International Nuclear Information System (INIS)

Xu, Wei; Wang, Ting; Xu, Shaoxin; Li, Fanghua; Deng, Chenguang; Wu, Lijun; Wu, Yuejin; Bian, Po

2016-01-01

Highlights: • Transcriptional gene silencing (TGS) in plants can be epigenetically alleviated by volatile signals from UV-C- irradiated neighboring plants. • Alleviation of TGS can be induced by UV-C irradiation through plant–plant–plant communication. • JA and SA signals take part in interplant communication for alleviation of TGS. - Abstract: Plant stress responses at the epigenetic level are expected to allow more permanent changes of gene expression and potentially long-term adaptation. While it has been reported that plants subjected to adverse environments initiate various stress responses in their neighboring plants, little is known regarding epigenetic responses to external stresses mediated by plant–plant communication. In this study, we show that DNA repetitive elements of Arabidopsis thaliana, whose expression is inhibited epigenetically by transcriptional gene silencing (TGS) mechanism, are activated by UV-C irradiation through airborne plant–plant and plant–plant–plant communications, accompanied by DNA demethylation at CHH sites. Moreover, the TGS is alleviated by direct treatments with exogenous methyl jasmonate (MeJA) and methyl salicylate (MeSA). Further, the plant–plant and plant–plant–plant communications are blocked by mutations in the biosynthesis or signaling of jasmonic acid (JA) or salicylic acid (SA), indicating that JA and SA pathways are involved in the interplant communication for epigenetic responses. For the plant–plant–plant communication, stress cues are relayed to the last set of receiver plants by promoting the production of JA and SA signals in relaying plants, which exhibit upregulated expression of genes for JA and SA biosynthesis and enhanced emanation of MeJA and MeSA.

Production of caffeoylmalic acid from glucose in engineered Escherichia coli.

Science.gov (United States)

Li, Tianzhen; Zhou, Wei; Bi, Huiping; Zhuang, Yibin; Zhang, Tongcun; Liu, Tao

2018-07-01

To achieve biosynthesis of caffeoylmalic acid from glucose in engineered Escherichia coli. We constructed the biosynthetic pathway of caffeoylmalic acid in E. coli by co-expression of heterologous genes RgTAL, HpaBC, At4CL2 and HCT2. To enhance the production of caffeoylmalic acid, we optimized the tyrosine metabolic pathway of E. coli to increase the supply of the substrate caffeic acid. Consequently, an E. coli-E. coli co-culture system was used for the efficient production of caffeoylmalic acid. The final titer of caffeoylmalic acid reached 570.1 mg/L. Microbial production of caffeoylmalic acid using glucose has application potential. In addition, microbial co-culture is an efficient tool for producing caffeic acid esters.

Analysis of the role of the Aspergillus niger aminolevulinic acid synthase (hemA) gene illustrates the difference between regulation of yeast and fungal haem- and sirohaem-dependent pathways

NARCIS (Netherlands)

Franken, A.C.; Christien Lokman, B.; Ram, A.F.; Hondel, C.A. van den; Weert, S. de; Punt, P.J.

2012-01-01

To increase knowledge on haem biosynthesis in filamentous fungi like Aspergillus niger, pathway-specific gene expression in response to haem and haem intermediates was analysed. This analysis showed that iron, 5′-aminolevulinic acid (ALA) and possibly haem control haem biosynthesis mostly via

Analysis of the role of the A. niger aminolevulinic acid synthase (hemA) gene illustrates the difference between regulation of yeast and fungal heme and siroheme dependent pathways

NARCIS (Netherlands)

A.F. Ram; C.A. van den Hondel; Christien Lokman; P.J. Punt; S. de Weert; A. Franken

2012-01-01

To increase knowledge on haem biosynthesis in filamentous fungi like Aspergillus niger, pathway-specific gene expression in response to haem and haem intermediates was analysed. This analysis showed that iron, 5'-aminolevulinic acid (ALA) and possibly haem control haem biosynthesis mostly via

Production of itaconic acid from acetate by engineering acid-tolerant Escherichia coli W.

Science.gov (United States)

Noh, Myung Hyun; Lim, Hyun Gyu; Woo, Sung Hwa; Song, Jinyi; Jung, Gyoo Yeol

2018-03-01

Utilization of abundant and cheap carbon sources can effectively reduce the production cost and enhance the economic feasibility. Acetate is a promising carbon source to achieve cost-effective microbial processes. In this study, we engineered an Escherichia coli strain to produce itaconic acid from acetate. As acetate is known to inhibit cell growth, we initially screened for a strain with a high tolerance to 10 g/L of acetate in the medium, and the W strain was selected as the host. Subsequently, the WC strain was obtained by overexpression of cad (encoding cis-aconitate decarboxylase) using a synthetic promoter and 5' UTR. However, the WC strain produced only 0.13 g/L itaconic acid because of low acetate uptake. To improve the production, the acetate assimilating pathway and glyoxylate shunt pathway were amplified by overexpression of pathway genes as well as its deregulation. The resulting strain, WCIAG4 produced 3.57 g/L itaconic acid (16.1% of theoretical maximum yield) after 88 hr of fermentation with rapid acetate assimilation. These efforts support that acetate can be a potential feedstock for biochemical production with engineered E. coli. © 2017 Wiley Periodicals, Inc.

Genomes of rumen bacteria encode atypical pathways for fermenting hexoses to short-chain fatty acids.

Science.gov (United States)

Hackmann, Timothy J; Ngugi, David Kamanda; Firkins, Jeffrey L; Tao, Junyi

2017-11-01

Bacteria have been thought to follow only a few well-recognized biochemical pathways when fermenting glucose or other hexoses. These pathways have been chiseled in the stone of textbooks for decades, with most sources rendering them as they appear in the classic 1986 text by Gottschalk. Still, it is unclear how broadly these pathways apply, given that they were established and delineated biochemically with only a few model organisms. Here, we show that well-recognized pathways often cannot explain fermentation products formed by bacteria. In the most extensive analysis of its kind, we reconstructed pathways for glucose fermentation from genomes of 48 species and subspecies of bacteria from one environment (the rumen). In total, 44% of these bacteria had atypical pathways, including several that are completely unprecedented for bacteria or any organism. In detail, 8% of bacteria had an atypical pathway for acetate formation; 21% of bacteria had an atypical pathway for propionate or succinate formation; 6% of bacteria had an atypical pathway for butyrate formation and 33% of bacteria had an atypical or incomplete Embden-Meyerhof-Parnas pathway. This study shows that reconstruction of metabolic pathways - a common goal of omics studies - could be incorrect if well-recognized pathways are used for reference. Furthermore, it calls for renewed efforts to delineate fermentation pathways biochemically. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

Alleviation of Drought Stress by Hydrogen Sulfide Is Partially Related to the Abscisic Acid Signaling Pathway in Wheat.

Science.gov (United States)

Ma, Dongyun; Ding, Huina; Wang, Chenyang; Qin, Haixia; Han, Qiaoxia; Hou, Junfeng; Lu, Hongfang; Xie, Yingxin; Guo, Tiancai

2016-01-01

Little information is available describing the effects of exogenous H2S on the ABA pathway in the acquisition of drought tolerance in wheat. In this study, we investigated the physiological parameters, the transcription levels of several genes involved in the abscisic acid (ABA) metabolism pathway, and the ABA and H2S contents in wheat leaves and roots under drought stress in response to exogenous NaHS treatment. The results showed that pretreatment with NaHS significantly increased plant height and the leaf relative water content of seedlings under drought stress. Compared with drought stress treatment alone, H2S application increased antioxidant enzyme activities and reduced MDA and H2O2 contents in both leaves and roots. NaHS pretreatment increased the expression levels of ABA biosynthesis and ABA reactivation genes in leaves; whereas the expression levels of ABA biosynthesis and ABA catabolism genes were up-regulated in roots. These results indicated that ABA participates in drought tolerance induced by exogenous H2S, and that the responses in leaves and roots are different. The transcription levels of genes encoding ABA receptors were up-regulated in response to NaHS pretreatment under drought conditions in both leaves and roots. Correspondingly, the H2S contents in leaves and roots were increased by NaHS pretreatment, while the ABA contents of leaves and roots decreased. This implied that there is complex crosstalk between these two signal molecules, and that the alleviation of drought stress by H2S, at least in part, involves the ABA signaling pathway.

Metabolic Engineering of the Shikimate Pathway for Production of Aromatics and Derived Compounds—Present and Future Strain Construction Strategies

Directory of Open Access Journals (Sweden)

Nils J. H. Averesch

2018-03-01

Full Text Available The aromatic nature of shikimate pathway intermediates gives rise to a wealth of potential bio-replacements for commonly fossil fuel-derived aromatics, as well as naturally produced secondary metabolites. Through metabolic engineering, the abundance of certain intermediates may be increased, while draining flux from other branches off the pathway. Often targets for genetic engineering lie beyond the shikimate pathway, altering flux deep in central metabolism. This has been extensively used to develop microbial production systems for a variety of compounds valuable in chemical industry, including aromatic and non-aromatic acids like muconic acid, para-hydroxybenzoic acid, and para-coumaric acid, as well as aminobenzoic acids and aromatic α-amino acids. Further, many natural products and secondary metabolites that are valuable in food- and pharma-industry are formed outgoing from shikimate pathway intermediates. (Reconstruction of such routes has been shown by de novo production of resveratrol, reticuline, opioids, and vanillin. In this review, strain construction strategies are compared across organisms and put into perspective with requirements by industry for commercial viability. Focus is put on enhancing flux to and through shikimate pathway, and engineering strategies are assessed in order to provide a guideline for future optimizations.

Vitamin C degradation products and pathways in the human lens.

Science.gov (United States)

Nemet, Ina; Monnier, Vincent M

2011-10-28

Vitamin C and its degradation products participate in chemical modifications of proteins in vivo through non-enzymatic glycation (Maillard reaction) and formation of different products called advanced glycation end products. Vitamin C levels are particularly high in selected tissues, such as lens, brain and adrenal gland, and its degradation products can inflict substantial protein damage via formation of advanced glycation end products. However, the pathways of in vivo vitamin C degradation are poorly understood. Here we have determined the levels of vitamin C oxidation and degradation products dehydroascorbic acid, 2,3-diketogulonic acid, 3-deoxythreosone, xylosone, and threosone in the human lens using o-phenylenediamine to trap both free and protein-bound adducts. In the protein-free fraction and water-soluble proteins (WSP), all five listed degradation products were identified. Dehydroascorbic acid, 2,3-diketogulonic acid, and 3-deoxythreosone were the major products in the protein-free fraction, whereas in the WSP, 3-deoxythreosone was the most abundant measured dicarbonyl. In addition, 3-deoxythreosone in WSP showed positive linear correlation with age (p degradation product bound to human lens proteins provides in vivo evidence for the non-oxidative pathway of dehydroascorbate degradation into erythrulose as a major pathway for vitamin C degradation in vivo.

Evolution of amino acid metabolism inferred through cladistic analysis.

Science.gov (United States)

Cunchillos, Chomin; Lecointre, Guillaume

2003-11-28

Because free amino acids were most probably available in primitive abiotic environments, their metabolism is likely to have provided some of the very first metabolic pathways of life. What were the first enzymatic reactions to emerge? A cladistic analysis of metabolic pathways of the 16 aliphatic amino acids and 2 portions of the Krebs cycle was performed using four criteria of homology. The analysis is not based on sequence comparisons but, rather, on coding similarities in enzyme properties. The properties used are shared specific enzymatic activity, shared enzymatic function without substrate specificity, shared coenzymes, and shared functional family. The tree shows that the earliest pathways to emerge are not portions of the Krebs cycle but metabolisms of aspartate, asparagine, glutamate, and glutamine. The views of Horowitz (Horowitz, N. H. (1945) Proc. Natl. Acad. Sci. U. S. A. 31, 153-157) and Cordón (Cordón, F. (1990) Tratado Evolucionista de Biologia, Aguilar, Madrid, Spain), according to which the upstream reactions in the catabolic pathways and the downstream reactions in the anabolic pathways are the earliest in evolution, are globally corroborated; however, with some exceptions. These are due to later opportunistic connections of pathways (actually already suggested by these authors). Earliest enzymatic functions are mostly catabolic; they were deaminations, transaminations, and decarboxylations. From the consensus tree we extracted four time spans for amino acid metabolism development. For some amino acids catabolism and biosynthesis occurred at the same time (Asp, Glu, Lys, Leu, Ala, Val, Ile, Pro, Arg). For others ultimate reactions that use amino acids as a substrate or as a product are distinct in time, with catabolism preceding anabolism for Asn, Gln, and Cys and anabolism preceding catabolism for Ser, Met, and Thr. Cladistic analysis of the structure of biochemical pathways makes hypotheses in biochemical evolution explicit and parsimonious.

Evaluation of anti-inflammatory effect of statins in chronic periodontitis

OpenAIRE

Snophia Suresh; Satya Narayana; P Jayakumar; Uma Sudhakar; V Pramod

2013-01-01

Objectives: Statins are the group of lipid-lowering drugs commonly used to control cardiovascular and cerebrovascular diseases. Statins have potential anti-inflammatory effect by blocking the intermediate metabolites of the mevalonate pathway. The objective of this study was to evaluate the anti-inflammatory effect of statin medication in chronic periodontitis patients. Materials and Methods: Thirty patients of age group between 40 and 60 years were selected from the outpatient pool of De...

The Role of ERK1/2 in the Progression of Anti-Androgen Resistance of mtDNA Deficient Prostate Cancer

Science.gov (United States)

2012-05-01

cancers are not viable if mitochondria as a whole are removed. Mitochondria serve multiple roles for the eukaryote including ATP synthesis , redox...transcriptional and/or translational regulation, PTMs, and/or proteasomal degradation by both sterol and non- sterol products of the mevalonate pathway (46, 47...intermediate in the synthesis of cholesterol. Cell Metab. 2005;1(3):179-89. 49. Nguyen AD, McDonald JG, Bruick RK, DeBose-Boyd RA. Hypoxia stimulates

Structure of active IspH enzyme from escherichia coli provides mechanistic insights into substrate reduction

KAUST Repository

Gräwert, Tobias

2009-07-20

The terminal step of the non-mevalonate pathway of terpene biosynthesis is catalyzed by IspH (see scheme). In the crystal structure of IspH from E. coli, a bound inorganic diphosphate ligand occupies the position of the diphosphate residue of the substrate. Together with mutation studies and theoretical calculations, these data support a mechanism which is analogous to the Birch reduction of allylic alcohols. © 2009 Wiley-VCH Verlag GmbH & Co. KGaA.

A splice donor mutation in NAA10 results in the dysregulation of the retinoic acid signaling pathway and causes Lenz microphthalmia syndrome

Science.gov (United States)

Esmailpour, Taraneh; Riazifar, Hamidreza; Liu, Linan; Donkervoort, Sandra; Huang, Vincent H; Madaan, Shreshtha; Shoucri, Bassem M; Busch, Anke; Wu, Jie; Towbin, Alexander; Chadwick, Robert B; Sequeira, Adolfo; Vawter, Marquis P; Sun, Guoli; Johnston, Jennifer J; Biesecker, Leslie G; Kawaguchi, Riki; Sun, Hui; Kimonis, Virginia; Huang, Taosheng

2014-01-01

Introduction Lenz microphthalmia syndrome (LMS) is a genetically heterogeneous X-linked disorder characterised by microphthalmia/anophthalmia, skeletal abnormalities, genitourinary malformations, and anomalies of the digits, ears, and teeth. Intellectual disability and seizure disorders are seen in about 60% of affected males. To date, no gene has been identified for LMS in the microphthalmia syndrome 1 locus (MCOPS1). In this study, we aim to find the disease-causing gene for this condition. Methods and results Using exome sequencing in a family with three affected brothers, we identified a mutation in the intron 7 splice donor site (c.471+2T→A) of the N-acetyltransferase NAA10 gene. NAA10 has been previously shown to be mutated in patients with Ogden syndrome, which is clinically distinct from LMS. Linkage studies for this family mapped the disease locus to Xq27-Xq28, which was consistent with the locus of NAA10. The mutation co-segregated with the phenotype and cDNA analysis showed aberrant transcripts. Patient fibroblasts lacked expression of full length NAA10 protein and displayed cell proliferation defects. Expression array studies showed significant dysregulation of genes associated with genetic forms of anophthalmia such as BMP4, STRA6, and downstream targets of BCOR and the canonical WNT pathway. In particular, STRA6 is a retinol binding protein receptor that mediates cellular uptake of retinol/vitamin A and plays a major role in regulating the retinoic acid signalling pathway. A retinol uptake assay showed that retinol uptake was decreased in patient cells. Conclusions We conclude that the NAA10 mutation is the cause of LMS in this family, likely through the dysregulation of the retinoic acid signalling pathway. PMID:24431331

Administration of Tauroursodeoxycholic Acid Attenuates Early Brain Injury via Akt Pathway Activation

Directory of Open Access Journals (Sweden)

Dongdong Sun

2017-07-01

Full Text Available Traumatic brain injury (TBI is one of the leading causes of trauma-induced mortality and disability, and emerging studies have shown that endoplasmic reticulum (ER stress plays an important role in the pathophysiology of TBI. Tauroursodeoxycholic acid (TUDCA, a hydrophilic bile acid, has been reported to act as an ER stress inhibitor and chemical chaperone and to have the potential to attenuate apoptosis and inflammation. To study the effects of TUDCA on brain injury, we subjected mice to TBI with a controlled cortical impact (CCI device. Using western blotting, we first examined TBI-induced changes in the expression levels of GRP78, an ER stress marker, p-PERK, PERK, p-eIF2a, eIF2a, ATF4, p-Akt, Akt, Pten, Bax, Bcl-2, Caspase-12 and CHOP, as well as changes in the mRNA levels of Akt, GRP78, Caspase-12 and CHOP using RT-PCR. Neuronal cell death was assessed by a terminal deoxynucleotidyl transferase (TdT-mediated dUTP nick end-labeling (TUNEL assay, and CHOP expression in neuronal cells was detected by double-immunofluorescence staining. Neurological and motor deficits were assessed by modified neurological severity scores (mNSS and beam balance and beam walking tests, and brain water content was also assessed. Our results indicated that ER stress peaked at 72 h after TBI and that TUDCA abolished ER stress and inhibited p-PERK, p-eIF2a, ATF4, Pten, Caspase-12 and CHOP expression levels. Moreover, our results show that TUDCA also improved neurological function and alleviated brain oedema. Additionally, TUDCA increased p-Akt expression and the Bcl-2/Bax ratio. However, the administration of the Akt inhibitor MK2206 or siRNA targeting of Akt abolished the beneficial effects of TUDCA. Taken together, our results indicate that TUDCA may attenuate early brain injury via Akt pathway activation.

Intestinal transport and metabolism of bile acids

Science.gov (United States)

Dawson, Paul A.; Karpen, Saul J.

2015-01-01

In addition to their classical roles as detergents to aid in the process of digestion, bile acids have been identified as important signaling molecules that function through various nuclear and G protein-coupled receptors to regulate a myriad of cellular and molecular functions across both metabolic and nonmetabolic pathways. Signaling via these pathways will vary depending on the tissue and the concentration and chemical structure of the bile acid species. Important determinants of the size and composition of the bile acid pool are their efficient enterohepatic recirculation, their host and microbial metabolism, and the homeostatic feedback mechanisms connecting hepatocytes, enterocytes, and the luminal microbiota. This review focuses on the mammalian intestine, discussing the physiology of bile acid transport, the metabolism of bile acids in the gut, and new developments in our understanding of how intestinal metabolism, particularly by the gut microbiota, affects bile acid signaling. PMID:25210150

Tributyltin and triphenyltin inhibit osteoclast differentiation through a retinoic acid receptor-dependent signaling pathway

International Nuclear Information System (INIS)

Yonezawa, Takayuki; Hasegawa, Shin-ichi; Ahn, Jae-Yong; Cha, Byung-Yoon; Teruya, Toshiaki; Hagiwara, Hiromi; Nagai, Kazuo; Woo, Je-Tae

2007-01-01

Organotin compounds, such as tributyltin (TBT) and triphenyltin (TPT), have been widely used in agriculture and industry. Although these compounds are known to have many toxic effects, including endocrine-disrupting effects, their effects on bone resorption are unknown. In this study, we investigated the effects of organotin compounds, such as monobutyltin (MBT), dibutyltin (DBT), TBT, and TPT, on osteoclast differentiation using mouse monocytic RAW264.7 cells. MBT and DBT had no effects, whereas TBT and TPT dose-dependently inhibited osteoclast differentiation at concentrations of 3-30 nM. Treatment with a retinoic acid receptor (RAR)-specific antagonist, Ro41-5253, restored the inhibition of osteoclastogenesis by TBT and TPT. TBT and TPT reduced receptor activator of nuclear factor-κB ligand (RANKL) induced nuclear factor of activated T cells (NFAT) c1 expression, and the reduction in NFATc1 expression was recovered by Ro41-5253. Our results suggest that TBT and TPT suppress osteoclastogenesis by inhibiting RANKL-induced NFATc1 expression via an RAR-dependent signaling pathway

Eicosapentaenoic acid (EPA) induced apoptosis in HepG2 cells through ROS–Ca{sup 2+}–JNK mitochondrial pathways

Energy Technology Data Exchange (ETDEWEB)

Zhang, Yuanyuan; Han, Lirong [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Qi, Wentao [Academy of State Administration of Grain, No.11 Baiwanzhuang Avenue, Xicheng District, Beijing, 100037 (China); Cheng, Dai; Ma, Xiaolei; Hou, Lihua [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Cao, Xiaohong, E-mail: caoxh@tust.edu.cn [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China); Wang, Chunling, E-mail: wangchunling@tust.edu.cn [Key Laboratory of Food Nutrition and Safety, Ministry of Education, College of Food Engineering and Biotechnology, Tianjin University of Science and Technology, No. 29, 13th Avenue, Tianjin Economy Technological Development Area, Tianjin 300457 (China)

2015-01-24

Highlights: • EPA evoked ROS formation, [Ca{sup 2+}]{sub c} accumulation, the opening of MPTP and the phosphorylation of JNK. • EPA-induced [Ca{sup 2+}]{sub c} elevation was depended on production of ROS. • EPA-induced ROS generation, [Ca{sup 2+}]{sub c} increase, and JNK activated caused MPTP opening. • The apoptosis induced by EPA was related to release of cytochrome C through the MPTP. • EPA induced HepG2 cells apoptosis through ROS–Ca{sup 2+}–JNK mitochondrial pathways. - Abstract: Eicosapentaenoic acid (EPA), a well-known dietary n−3 PUFAS, has been considered to inhibit proliferation of tumor cells. However, the molecular mechanism related to EPA-induced liver cancer cells apoptosis has not been reported. In this study, we investigated the effect of EPA on HepG2 cells proliferation and apoptosis mechanism through mitochondrial pathways. EPA inhibited proliferation of HepG2 cells in a dose-dependent manner and had no significant effect on the cell viability of humor normal liver L-02 cells. It was found that EPA initially evoked ROS formation, leading to [Ca{sup 2+}]{sub c} accumulation and the mitochondrial permeability transition pore (MPTP) opening; EPA-induced HepG2 cells apoptosis was inhibited by N-acetylcysteine (NAC, an inhibitor of ROS), 1,2-bis (2-aminophenoxy) ethane-N,N,N′,N′-tetraacetic acid (BAPTA-AM, a chelator of calcium) and CsA (inhibitor of MPTP). The relationship between ROS production, the increase of cytoplasmic Ca and MPTP opening was detected. It seems that ROS may act as an upstream regulator of EPA-induced [Ca{sup 2+}]{sub c} generation, moreover, generation of ROS, overload of mitochondrial [Ca{sup 2+}]{sub c}, and JNK activated cause the opening of MPTP. Western blotting results showed that EPA elevated the phosphorylation status of JNK, processes associated with the ROS generation. Simultaneously, the apoptosis induced by EPA was related to release of cytochrome C from mitochondria to cytoplasm through the MPTP

Salvianolic acid A reverses cisplatin resistance in lung cancer A549 cells by targeting c-met and attenuating Akt/mTOR pathway

Directory of Open Access Journals (Sweden)

Xia-li Tang

2017-09-01

Full Text Available Drug resistance is one of the leading causes of chemotherapy failure in non-small cell lung cancer (NSCLC treatment. The purpose of this study was to investigate the role of c-met in human lung cancer cisplatin resistance cell line (A549/DDP and the reversal mechanism of salvianolic acid A (SAA, a phenolic active compound extracted from Salvia miltiorrhiza. In this study, we found that A549/DDP cells exert up-regulation of c-met by activating the Akt/mTOR signaling pathway. We also show that SAA could increase the chemotherapeutic efficacy of cisplatin, suggesting a synergistic effect of SAA and cisplatin. Moreover, we revealed that SAA enhanced sensitivity to cisplatin in A549/DDP cells mainly through suppression of the c-met/AKT/mTOR signaling pathway. Knockdown of c-met revealed similar effects as that of SAA in A549/DDP cells. In addition, SAA effectively prevented multidrug resistance associated protein1 (MDR1 up-regulation in A549/DDP cells. Taken together, our results indicated that SAA suppressed c-met expression and enhanced the sensitivity of lung adenocarcinoma A549 cells to cisplatin through AKT/mTOR signaling pathway.

A Functional Tricarboxylic Acid Cycle Operates during Growth of Bordetella pertussis on Amino Acid Mixtures as Sole Carbon Substrates.

Directory of Open Access Journals (Sweden)

Marie Izac

Full Text Available It has been claimed that citrate synthase, aconitase and isocitrate dehydrogenase activities are non-functional in Bordetella pertussis and that this might explain why this bacterium's growth is sometimes associated with accumulation of polyhydroxybutyrate (PHB and/or free fatty acids. However, the sequenced genome includes the entire citric acid pathway genes. Furthermore, these genes were expressed and the corresponding enzyme activities detected at high levels for the pathway when grown on a defined medium imitating the amino acid content of complex media often used for growth of this pathogenic microorganism. In addition, no significant PHB or fatty acids could be detected. Analysis of the carbon balance and stoichiometric flux analysis based on specific rates of amino acid consumption, and estimated biomass requirements coherent with the observed growth rate, clearly indicate that a fully functional tricarboxylic acid cycle operates in contrast to previous reports.

Comparative Transcriptome Analysis Reveals the Influence of Abscisic Acid on the Metabolism of Pigments, Ascorbic Acid and Folic Acid during Strawberry Fruit Ripening.

Directory of Open Access Journals (Sweden)

Dongdong Li

Full Text Available A comprehensive investigation of abscisic acid (ABA biosynthesis and its influence on other important phytochemicals is critical for understanding the versatile roles that ABA plays during strawberry fruit ripening. Using RNA-seq technology, we sampled strawberry fruit in response to ABA or nordihydroguaiaretic acid (NDGA; an ABA biosynthesis blocker treatment during ripening and assessed the expression changes of genes involved in the metabolism of pigments, ascorbic acid (AsA and folic acid in the receptacles. The transcriptome analysis identified a lot of genes differentially expressed in response to ABA or NDGA treatment. In particular, genes in the anthocyanin biosynthesis pathway were actively regulated by ABA, with the exception of the gene encoding cinnamate 4-hydroxylase. Chlorophyll degradation was accelerated by ABA mainly owing to the higher expression of gene encoding pheide a oxygenase. The decrease of β-carotene content was accelerated by ABA treatment and delayed by NDGA. A high negative correlation rate was found between ABA and β-carotene content, indicating the importance of the requirement for ABA synthesis during fruit ripening. In addition, evaluation on the folate biosynthetic pathway indicate that ABA might have minor function in this nutrient's biosynthesis process, however, it might be involved in its homeostasis. Surprisingly, though AsA content accumulated during fruit ripening, expressions of genes involved in its biosynthesis in the receptacles were significantly lower in ABA-treated fruits. This transcriptome analysis expands our understanding of ABA's role in phytochemical metabolism during strawberry fruit ripening and the regulatory mechanisms of ABA on these pathways were discussed. Our study provides a wealth of genetic information in the metabolism pathways and may be helpful for molecular manipulation in the future.

Genomes of rumen bacteria encode atypical pathways for fermenting hexoses to short-chain fatty acids

KAUST Repository

Hackmann, Timothy J.

2017-09-11

Bacteria have been thought to follow only a few well-recognized biochemical pathways when fermenting glucose or other hexoses. These pathways have been chiseled in the stone of textbooks for decades, with most sources rendering them as they appear in the classic 1986 text by Gottschalk. Still, it is unclear how broadly these pathways apply, given that they were established and delineated biochemically with only a few model organisms. Here we show that well-recognized pathways often cannot explain fermentation products formed by bacteria. In the most extensive analysis of its kind, we reconstructed pathways for glucose fermentation from genomes of 48 species and subspecies of bacteria from one environment (the rumen). In total, 44% of these bacteria had atypical pathways, including several that are completely unprecedented for bacteria or any organism. In detail, 8% of bacteria had an atypical pathway for acetate formation; 21% for propionate or succinate formation; 6% for butyrate formation; and 33% had an atypical or incomplete Embden-Meyerhof-Parnas pathway. This study shows that reconstruction of metabolic pathways-a common goal of omics studies-could be incorrect if well-recognized pathways are used for reference. Further, it calls for renewed efforts to delineate fermentation pathways biochemically. This article is protected by copyright. All rights reserved.

Theoretical Study on Free Fatty Acid Elimination Mechanism for Waste Cooking Oils to Biodiesel over Acid Catalyst.

Science.gov (United States)

Wang, Kai; Zhang, Xiaochao; Zhang, Jilong; Zhang, Zhiqiang; Fan, Caimei; Han, Peide

2016-05-01

A theoretical investigation on the esterification mechanism of free fatty acid (FFA) in waste cooking oils (WCOs) has been carried out using DMol(3) module based on the density functional theory (DFT). Three potential pathways of FFA esterification reaction are designed to achieve the formation of fatty acid methyl ester (FAME), and calculated results show that the energy barrier can be efficiently reduced from 88.597kcal/mol to 15.318kcal/mol by acid catalyst. The molar enthalpy changes (ΔrHm°) of designed pathways are negative, indicating that FFA esterification reaction is an exothermic process. The obtained favorable energy pathway is: H(+) firstly activates FFA, then the intermediate combines with methanol to form a tetrahedral structure, and finally, producing FAME after removing a water molecule. The rate-determining step is the combination of the activated FFA with methanol, and the activation energy is about 11.513kcal/mol at 298.15K. Our results should provide basic and reliable theoretical data for further understanding the elimination mechanism of FFA over acid catalyst in the conversion of WCOs to biodiesel products. Copyright © 2016 Elsevier Inc. All rights reserved.

Metabolic solutions to the biosynthesis of some diaminomonocarboxylic acids in nature: Formation in cyanobacteria of the neurotoxins 3-N-methyl-2,3-diaminopropanoic acid (BMAA) and 2,4-diaminobutanoic acid (2,4-DAB).

Science.gov (United States)

Nunn, Peter B; Codd, Geoffrey A

2017-12-01

The non-encoded diaminomonocarboxylic acids, 3-N-methyl-2,3-diaminopropanoic acid (syn: α-amino-β-methylaminopropionic acid, MeDAP; β-N-methylaminoalanine, BMAA) and 2,4-diaminobutanoic acid (2,4-DAB), are distributed widely in cyanobacterial species in free and bound forms. Both amino acids are neurotoxic in whole animal and cell-based bioassays. The biosynthetic pathway to 2,4-DAB is well documented in bacteria and in one higher plant species, but has not been confirmed in cyanobacteria. The biosynthetic pathway to BMAA is unknown. This review considers possible metabolic routes, by analogy with reactions used in other species, by which these amino acids might be biosynthesised by cyanobacteria, which are a widespread potential environmental source of these neurotoxins. Where possible, the gene expression that might be implicated in these biosyntheses is discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

The long and winding road: transport pathways for amino acids in Arabidopsis seeds.

Science.gov (United States)

Karmann, Julia; Müller, Benedikt; Hammes, Ulrich Z

2018-03-16

Pathways for assimilates. During their life cycle, plants alternate between a haploid stage, the gametophyte, and a diploid stage, the sporophyte. In higher plants, meiosis generates the gametophyte deeply embedded in the maternal tissue of the flower. The megaspore mother cell undergoes meiosis, and then, the surviving megaspore of the four megaspores produced undergoes mitotic divisions and finally gives rise to the female gametophyte, consisting of the egg cell, two synergids, the central cell, which due to the fusion of two nuclei is diploid (double haploid) in Arabidopsis and most angiosperms and the antipods, whose number is not fixed and varies significantly between species (Yadegari and Drews in Plant Cell 16(Suppl):S133-S141, 2004). The maternal tissues that harbor the female gametophyte and the female gametophyte are referred to as the ovule (Fig. 1). Double fertilization of the egg cell and the central cell by the two generative nuclei of the pollen leads to the diploid embryo and the endosperm, respectively (Hamamura et al. in Curr Opin Plant Biol 15:70-77, 2012). Upon fertilization, the ovule is referred to as the seed. Seeds combine two purposes: to harbor storage compounds for use by the embryo upon germination and to protect the embryo until the correct conditions for germination are encountered. As a consequence, seeds are the plant tissue that is of highest nutritional value and the human diet, by a considerable amount, consists of seeds or seed-derived products. Amino acids are of special interest, because plants serve as the main source for the so-called essential amino acids, that animals cannot synthesize de novo and are therefore often a limiting factor for human growth and development (WHO in Protein and amino acid requirements in human nutrition. WHO technical report series, WHO, Geneva, 2007). The plant embryo needs amino acids for general protein synthesis, and additionally they are used to synthesize storage proteins in the seeds of

Anti-Inflammatory Effects of Benfotiamine are Mediated Through the Regulation of Arachidonic Acid Pathway in Macrophages

Science.gov (United States)

Shoeb, Mohammad; Ramana, Kota V

2011-01-01

Benfotiamine, a lipid-soluble analogue of vitamin B1, is a potent anti-oxidant that is used as a food supplement for the treatment of diabetic complications. Our recent study indicates a novel role of benfotiamine in the prevention of bacterial endotoxin, lipopolysaccharide (LPS)-induced cytotoxicity and inflammatory response in murine macrophages. Nevertheless, it remains unclear how benfotiamine mediates anti-inflammatory effects. In this study, we investigated the anti-inflammatory role of benfotiamine in regulating the arachidonic acid (AA) pathway generated inflammatory lipid mediators in RAW 264.7 macrophages. Benfotiamine prevented the LPS-induced activation of cPLA2 and release of AA metabolites such as leukotrienes (LTB4), prostaglandin E2 (PGE2), thromboxanes 2 (TXB2) and prostacyclin (PGI2) in macrophages. Further, LPS-induced expressions of AA metabolizing enzymes such as COX-2, LOX-5, TXB synthase and PGI2 synthase were significantly blocked by benfotiamine. Furthermore, benfotiamine prevented the LPS-induced phosphorylation of ERK1/2 and expression of transcription factors NF-kB, and Egr-1. Benfotiamine also prevented the LPS-induced oxidative stress and protein-HNE adducts formation. Most importantly, as compared to specific COX-2 and LOX-5 inhibitors, benfotiamine significantly prevented the LPS-induced macrophage death and monocytes adhesion to endothelial cells. Thus, our studies indicate that the dual regulation of COX and LOX pathways in AA metabolism could be a novel mechanism by which benfotiamine exhibits its potential anti-inflammatory response. PMID:22067901

Resistance to ursodeoxycholic acid-induced growth arrest can also result in resistance to deoxycholic acid-induced apoptosis and increased tumorgenicity

International Nuclear Information System (INIS)

Powell, Ashley A; Akare, Sandeep; Qi, Wenqing; Herzer, Pascal; Jean-Louis, Samira; Feldman, Rebecca A; Martinez, Jesse D

2006-01-01

There is a large body of evidence which suggests that bile acids increase the risk of colon cancer and act as tumor promoters, however, the mechanism(s) of bile acids mediated tumorigenesis is not clear. Previously we showed that deoxycholic acid (DCA), a tumorogenic bile acid, and ursodeoxycholic acid (UDCA), a putative chemopreventive agent, exhibited distinct biological effects, yet appeared to act on some of the same signaling molecules. The present study was carried out to determine whether there is overlap in signaling pathways activated by tumorogenic bile acid DCA and chemopreventive bile acid UDCA. To determine whether there was an overlap in activation of signaling pathways by DCA and UDCA, we mutagenized HCT116 cells and then isolated cell lines resistant to UDCA induced growth arrest. These lines were then tested for their response to DCA induced apoptosis. We found that a majority of the cell lines resistant to UDCA-induced growth arrest were also resistant to DCA-induced apoptosis, implying an overlap in DCA and UDCA mediated signaling. Moreover, the cell lines which were the most resistant to DCA-induced apoptosis also exhibited a greater capacity for anchorage independent growth. We conclude that UDCA and DCA have overlapping signaling activities and that disregulation of these pathways can lead to a more advanced neoplastic phenotype

Amino Acid Crossword Puzzle

Science.gov (United States)

Sims, Paul A.

2011-01-01

Learning the 20 standard amino acids is an essential component of an introductory course in biochemistry. Later in the course, the students study metabolism and learn about various catabolic and anabolic pathways involving amino acids. Learning new material or concepts often is easier if one can connect the new material to what one already knows;…

Further increased production of free fatty acids by overexpressing a predicted transketolase gene of the pentose phosphate pathway in Aspergillus oryzae faaA disruptant.

Science.gov (United States)

Tamano, Koichi; Miura, Ai

2016-09-01

Free fatty acids are useful as source materials for the production of biodiesel fuel and various chemicals such as pharmaceuticals and dietary supplements. Previously, we attained a 9.2-fold increase in free fatty acid productivity by disrupting a predicted acyl-CoA synthetase gene (faaA, AO090011000642) in Aspergillus oryzae. In this study, we achieved further increase in the productivity by overexpressing a predicted transketolase gene of the pentose phosphate pathway in the faaA disruptant. The A. oryzae genome is predicted to have three transketolase genes and overexpression of AO090023000345, one of the three genes, resulted in phenotypic change and further increase (corresponding to an increased production of 0.38 mmol/g dry cell weight) in free fatty acids at 1.4-fold compared to the faaA disruptant. Additionally, the biomass of hyphae increased at 1.2-fold by the overexpression. As a result, free fatty acid production yield per liter of liquid culture increased at 1.7-fold by the overexpression.

Linking γ-aminobutyric acid A receptor to epidermal growth factor receptor pathways activation in human prostate cancer.

Science.gov (United States)

Wu, Weijuan; Yang, Qing; Fung, Kar-Ming; Humphreys, Mitchell R; Brame, Lacy S; Cao, Amy; Fang, Yu-Ting; Shih, Pin-Tsen; Kropp, Bradley P; Lin, Hsueh-Kung

2014-03-05

Neuroendocrine (NE) differentiation has been attributed to the progression of castration-resistant prostate cancer (CRPC). Growth factor pathways including the epidermal growth factor receptor (EGFR) signaling have been implicated in the development of NE features and progression to a castration-resistant phenotype. However, upstream molecules that regulate the growth factor pathway remain largely unknown. Using androgen-insensitive bone metastasis PC-3 cells and androgen-sensitive lymph node metastasis LNCaP cells derived from human prostate cancer (PCa) patients, we demonstrated that γ-aminobutyric acid A receptor (GABA(A)R) ligand (GABA) and agonist (isoguvacine) stimulate cell proliferation, enhance EGF family members expression, and activate EGFR and a downstream signaling molecule, Src, in both PC-3 and LNCaP cells. Inclusion of a GABA(A)R antagonist, picrotoxin, or an EGFR tyrosine kinase inhibitor, Gefitinib (ZD1839 or Iressa), blocked isoguvacine and GABA-stimulated cell growth, trans-phospohorylation of EGFR, and tyrosyl phosphorylation of Src in both PCa cell lines. Spatial distributions of GABAAR α₁ and phosphorylated Src (Tyr416) were studied in human prostate tissues by immunohistochemistry. In contrast to extremely low or absence of GABA(A)R α₁-positive immunoreactivity in normal prostate epithelium, elevated GABA(A)R α₁ immunoreactivity was detected in prostate carcinomatous glands. Similarly, immunoreactivity of phospho-Src (Tyr416) was specifically localized and limited to the nucleoli of all invasive prostate carcinoma cells, but negative in normal tissues. Strong GABAAR α₁ immunoreactivity was spatially adjacent to the neoplastic glands where strong phospho-Src (Tyr416)-positive immunoreactivity was demonstrated, but not in adjacent to normal glands. These results suggest that the GABA signaling is linked to the EGFR pathway and may work through autocrine or paracine mechanism to promote CRPC progression. Copyright © 2013 Elsevier

Cocrystals of a 1,2,4-thiadiazole-based potent neuroprotector with gallic acid: solubility, thermodynamic stability relationships and formation pathways.

Science.gov (United States)

Surov, Artem O; Churakov, Andrei V; Proshin, Alexey N; Dai, Xia-Lin; Lu, Tongbu; Perlovich, German L

2018-05-30

Three distinct solid forms, namely anhydrous cocrystals with 2 : 1 and 1 : 1 drug/acid ratios ([TDZ : GA] (2 : 1), [TDZ : GA] (1 : 1)), and a hydrated one having 1 : 1 : 1 drug/acid/water stoichiometry ([TDZ : GA : H2O] (1 : 1 : 1)), have been formed by cocrystallization of the biologically active 1,2,4-thiadiazole derivative (TDZ) with gallic acid (GA). The thermodynamic stability relationships between the cocrystals were rationalized in terms of Gibbs energies of the formation reactions and further verified by performing a set of competitive and exchange mechanochemical reactions. Interestingly, competitive grinding in the presence of the structurally related vanillic acid led to the formation of a new polymorphic form of the [TDZ : Vanillic acid] (1 : 1) cocrystal, which was promoted by gallic acid. The mechanochemical method was also applied to elucidate the alternative pathways of the [TDZ : GA : H2O] (1 : 1 : 1) cocrystal formation. Direct cocrystallization of TDZ with GA monohydrate was found to proceed much faster than the reaction of TDZ and anhydrous GA in the presence of an acetonitrile/water mixture, which may indicate the presence of a transitional stage. According to dissolution studies, the [TDZ : GA : H2O] (1 : 1 : 1) cocrystal was ca. 6.6 times more soluble than the parent 1,2,4-thiadiazole at pH 2.0 and 25.0 °C. The apparent two-step dehydration behavior of the [TDZ : GA : H2O] (1 : 1 : 1) cocrystal monohydrate was clarified by analyzing the intermolecular interactions of water molecules with the crystalline environment derived from solid state DFT calculations.

Proteomic analysis of lettuce seed germination and thermoinhibition by sampling of individual seeds at germination and removal of storage proteins by polyethylene glycol fractionation.

Science.gov (United States)

Wang, Wei-Qing; Song, Bin-Yan; Deng, Zhi-Jun; Wang, Yue; Liu, Shu-Jun; Møller, Ian Max; Song, Song-Quan

2015-04-01

Germination and thermoinhibition in lettuce (Lactuca sativa 'Jianyexianfeng No. 1') seeds were investigated by a proteomic comparison among dry seeds, germinated seeds at 15°C, at 15°C after imbibition at 25°C for 48 h, or at 25°C in KNO3 (all sampled individually at germination), and ungerminated seeds at 25°C, a thermoinhibitory temperature. Before two-dimensional gel electrophoresis analysis, storage proteins (greater than 50% of total extractable protein) were removed by polyethylene glycol precipitation, which significantly improved the detection of less abundant proteins on two-dimensional gels. A total of 108 protein spots were identified to change more than 2-fold (Pseeds than in ungerminated 25°C seeds. Gene expression of 12 of those proteins correlated well with the protein accumulation. Methionine metabolism, ethylene production, lipid mobilization, cell elongation, and detoxification of aldehydes were revealed to be potentially related to lettuce seed germination and thermoinhibition. Accumulation of three proteins and expression of five genes participating in the mevalonate (MVA) pathway of isoprenoid biosynthesis correlated positively with seed germinability. Inhibition of this pathway by lovastatin delayed seed germination and increased the sensitivity of germination to abscisic acid. MVA pathway-derived products, cytokinins, partially reversed the lovastatin inhibition of germination and released seed thermoinhibition at 25°C. We conclude that the MVA pathway for isoprenoid biosynthesis is involved in lettuce seed germination and thermoinhibition. © 2015 American Society of Plant Biologists. All Rights Reserved.

The effect of selected metals on the central metabolic pathways in ...

African Journals Online (AJOL)

compounds, interfere with xenobiotic metabolic pathways, and may also affect glycolysis, the Krebs cycle, oxidative phosphorylation, protein amino acid metabolism as well as carbohydrate and lipid metabolism. Therefore, in this review, we discuss the two phases of the central metabolic pathways, as well as how metals ...

Up-regulation of an N-terminal truncated 3-hydroxy-3-methylglutaryl CoA reductase enhances production of essential oils and sterols in transgenic Lavandula latifolia.

Science.gov (United States)

Muñoz-Bertomeu, Jesús; Sales, Ester; Ros, Roc; Arrillaga, Isabel; Segura, Juan

2007-11-01

Spike lavender (Lavandula latifolia) essential oil is widely used in the perfume, cosmetic, flavouring and pharmaceutical industries. Thus, modifications of yield and composition of this essential oil by genetic engineering should have important scientific and commercial applications. We generated transgenic spike lavender plants expressing the Arabidopsis thaliana HMG1 cDNA, encoding the catalytic domain of 3-hydroxy-3-methylglutaryl CoA reductase (HMGR1S), a key enzyme of the mevalonic acid (MVA) pathway. Transgenic T0 plants accumulated significantly more essential oil constituents as compared to controls (up to 2.1- and 1.8-fold in leaves and flowers, respectively). Enhanced expression of HMGR1S also increased the amount of the end-product sterols, beta-sitosterol and stigmasterol (average differences of 1.8- and 1.9-fold, respectively), but did not affect the accumulation of carotenoids or chlorophylls. We also analysed T1 plants derived from self-pollinated seeds of T0 lines that flowered after growing for 2 years in the greenhouse. The increased levels of essential oil and sterols observed in the transgenic T0 plants were maintained in the progeny that inherited the HMG1 transgene. Our results demonstrate that genetic manipulation of the MVA pathway increases essential oil yield in spike lavender, suggesting a contribution for this cytosolic pathway to monoterpene and sesquiterpene biosynthesis in leaves and flowers of the species.

Modulation of neurotrophic signaling pathways by polyphenols

Directory of Open Access Journals (Sweden)

Moosavi F

2015-12-01

Full Text Available Fatemeh Moosavi,1,2 Razieh Hosseini,1,2 Luciano Saso,3 Omidreza Firuzi1 1Medicinal and Natural Products Chemistry Research Center, Shiraz University of Medical Sciences, Shiraz, Iran; 2Department of Pharmacology, School of Veterinary Medicine, Shiraz University, Shiraz, Iran; 3Department of Physiology and Pharmacology “Vittorio Erspamer”, Sapienza University of Rome, Rome, Italy Abstract: Polyphenols are an important class of phytochemicals, and several lines of evidence have demonstrated their beneficial effects in the context of a number of pathologies including neurodegenerative disorders such as Alzheimer’s and Parkinson’s disease. In this report, we review the studies on the effects of polyphenols on neuronal survival, growth, proliferation and differentiation, and the signaling pathways involved in these neurotrophic actions. Several polyphenols including flavonoids such as baicalein, daidzein, luteolin, and nobiletin as well as nonflavonoid polyphenols such as auraptene, carnosic acid, curcuminoids, and hydroxycinnamic acid derivatives including caffeic acid phentyl ester enhance neuronal survival and promote neurite outgrowth in vitro, a hallmark of neuronal differentiation. Assessment of underlying mechanisms, especially in PC12 neuronal-like cells, reveals that direct agonistic effect on tropomyosin receptor kinase (Trk receptors, the main receptors of neurotrophic factors including nerve growth factor (NGF and brain-derived neurotrophic factor (BDNF explains the action of few polyphenols such as 7,8-dihydroxyflavone. However, several other polyphenolic compounds activate extracellular signal-regulated kinase (ERK and phosphoinositide 3-kinase (PI3K/Akt pathways. Increased expression of neurotrophic factors in vitro and in vivo is the mechanism of neurotrophic action of flavonoids such as scutellarin, daidzein, genistein, and fisetin, while compounds like apigenin and ferulic acid increase cyclic adenosine monophosphate

Metabolic engineering of Pediococcus acidilactici BD16 for production of vanillin through ferulic acid catabolic pathway and process optimization using response surface methodology.

Science.gov (United States)

Kaur, Baljinder; Chakraborty, Debkumar; Kumar, Balvir

2014-10-01

Occurrence of feruloyl-CoA synthetase (fcs) and enoyl-CoA hydratase (ech) genes responsible for the bioconversion of ferulic acid to vanillin have been reported and characterized from Amycolatopsis sp., Streptomyces sp., and Pseudomonas sp. Attempts have been made to express these genes in Escherichia coli DH5α, E. coli JM109, and Pseudomonas fluorescens. However, none of the lactic acid bacteria strain having GRAS status was previously proposed for heterologous expression of fcs and ech genes for production of vanillin through biotechnological process. Present study reports heterologous expression of vanillin synthetic gene cassette bearing fcs and ech genes in a dairy isolate Pediococcus acidilactici BD16. After metabolic engineering, statistical optimization of process parameters that influence ferulic acid to vanillin biotransformation in the recombinant strain was carried out using central composite design of response surface methodology. After scale-up of the process, 3.14 mM vanillin was recovered from 1.08 mM ferulic acid per milligram of recombinant cell biomass within 20 min of biotransformation. From LCMS-ESI spectral analysis, a metabolic pathway of phenolic biotransformations was predicted in the recombinant P. acidilactici BD16 (fcs (+)/ech (+)).

13C based proteinogenic amino acid (PAA) and metabolic flux ratio analysis of Lactococcus lactis reveals changes in pentose phosphate (PP) pathway in response to agitation and temperature related stresses.

Science.gov (United States)

Azizan, Kamalrul Azlan; Ressom, Habtom W; Mendoza, Eduardo R; Baharum, Syarul Nataqain

2017-01-01

Lactococcus lactis subsp. cremoris MG1363 is an important starter culture for dairy fermentation. During industrial fermentations, L. lactis is constantly exposed to stresses that affect the growth and performance of the bacterium. Although the response of L. lactis to several stresses has been described, the adaptation mechanisms at the level of in vivo fluxes have seldom been described. To gain insights into cellular metabolism, 13 C metabolic flux analysis and gas chromatography mass spectrometry (GC-MS) were used to measure the flux ratios of active pathways in the central metabolism of L. lactis when subjected to three conditions varying in temperature (30°C, 37°C) and agitation (with and without agitation at 150 rpm). Collectively, the concentrations of proteinogenic amino acids (PAAs) and free fatty acids (FAAs) were compared, and Pearson correlation analysis ( r ) was calculated to measure the pairwise relationship between PAAs. Branched chain and aromatic amino acids, threonine, serine, lysine and histidine were correlated strongly, suggesting changes in flux regulation in glycolysis, the pentose phosphate (PP) pathway, malic enzyme and anaplerotic reaction catalysed by pyruvate carboxylase (pycA). Flux ratio analysis revealed that glucose was mainly converted by glycolysis, highlighting the stability of L. lactis' central carbon metabolism despite different conditions. Higher flux ratios through oxaloacetate (OAA) from pyruvate (PYR) reaction in all conditions suggested the activation of pyruvate carboxylate (pycA) in L. lactis , in response to acid stress during exponential phase. Subsequently, more significant flux ratio differences were seen through the oxidative and non-oxidative pentose phosphate (PP) pathways, malic enzyme, and serine and C1 metabolism, suggesting NADPH requirements in response to environmental stimuli. These reactions could play an important role in optimization strategies for metabolic engineering in L. lactis . Overall, the

13C based proteinogenic amino acid (PAA and metabolic flux ratio analysis of Lactococcus lactis reveals changes in pentose phosphate (PP pathway in response to agitation and temperature related stresses

Directory of Open Access Journals (Sweden)

Kamalrul Azlan Azizan

2017-07-01

Full Text Available Lactococcus lactis subsp. cremoris MG1363 is an important starter culture for dairy fermentation. During industrial fermentations, L. lactis is constantly exposed to stresses that affect the growth and performance of the bacterium. Although the response of L. lactis to several stresses has been described, the adaptation mechanisms at the level of in vivo fluxes have seldom been described. To gain insights into cellular metabolism, 13C metabolic flux analysis and gas chromatography mass spectrometry (GC-MS were used to measure the flux ratios of active pathways in the central metabolism of L. lactis when subjected to three conditions varying in temperature (30°C, 37°C and agitation (with and without agitation at 150 rpm. Collectively, the concentrations of proteinogenic amino acids (PAAs and free fatty acids (FAAs were compared, and Pearson correlation analysis (r was calculated to measure the pairwise relationship between PAAs. Branched chain and aromatic amino acids, threonine, serine, lysine and histidine were correlated strongly, suggesting changes in flux regulation in glycolysis, the pentose phosphate (PP pathway, malic enzyme and anaplerotic reaction catalysed by pyruvate carboxylase (pycA. Flux ratio analysis revealed that glucose was mainly converted by glycolysis, highlighting the stability of L. lactis’ central carbon metabolism despite different conditions. Higher flux ratios through oxaloacetate (OAA from pyruvate (PYR reaction in all conditions suggested the activation of pyruvate carboxylate (pycA in L. lactis, in response to acid stress during exponential phase. Subsequently, more significant flux ratio differences were seen through the oxidative and non-oxidative pentose phosphate (PP pathways, malic enzyme, and serine and C1 metabolism, suggesting NADPH requirements in response to environmental stimuli. These reactions could play an important role in optimization strategies for metabolic engineering in L. lactis. Overall

13C based proteinogenic amino acid (PAA) and metabolic flux ratio analysis of Lactococcus lactis reveals changes in pentose phosphate (PP) pathway in response to agitation and temperature related stresses

Science.gov (United States)

2017-01-01

Lactococcus lactis subsp. cremoris MG1363 is an important starter culture for dairy fermentation. During industrial fermentations, L. lactis is constantly exposed to stresses that affect the growth and performance of the bacterium. Although the response of L. lactis to several stresses has been described, the adaptation mechanisms at the level of in vivo fluxes have seldom been described. To gain insights into cellular metabolism, 13C metabolic flux analysis and gas chromatography mass spectrometry (GC-MS) were used to measure the flux ratios of active pathways in the central metabolism of L. lactis when subjected to three conditions varying in temperature (30°C, 37°C) and agitation (with and without agitation at 150 rpm). Collectively, the concentrations of proteinogenic amino acids (PAAs) and free fatty acids (FAAs) were compared, and Pearson correlation analysis (r) was calculated to measure the pairwise relationship between PAAs. Branched chain and aromatic amino acids, threonine, serine, lysine and histidine were correlated strongly, suggesting changes in flux regulation in glycolysis, the pentose phosphate (PP) pathway, malic enzyme and anaplerotic reaction catalysed by pyruvate carboxylase (pycA). Flux ratio analysis revealed that glucose was mainly converted by glycolysis, highlighting the stability of L. lactis’ central carbon metabolism despite different conditions. Higher flux ratios through oxaloacetate (OAA) from pyruvate (PYR) reaction in all conditions suggested the activation of pyruvate carboxylate (pycA) in L. lactis, in response to acid stress during exponential phase. Subsequently, more significant flux ratio differences were seen through the oxidative and non-oxidative pentose phosphate (PP) pathways, malic enzyme, and serine and C1 metabolism, suggesting NADPH requirements in response to environmental stimuli. These reactions could play an important role in optimization strategies for metabolic engineering in L. lactis. Overall, the

Hyperthermophilic Archaeon Thermococcus kodakarensis Utilizes a Four-Step Pathway for NAD+ Salvage through Nicotinamide Deamination.

Science.gov (United States)

Hachisuka, Shin-Ichi; Sato, Takaaki; Atomi, Haruyuki

2018-06-01

Many organisms possess pathways that regenerate NAD + from its degradation products, and two pathways are known to salvage NAD + from nicotinamide (Nm). One is a four-step pathway that proceeds through deamination of Nm to nicotinic acid (Na) by Nm deamidase and phosphoribosylation to nicotinic acid mononucleotide (NaMN), followed by adenylylation and amidation. Another is a two-step pathway that does not involve deamination and directly proceeds with the phosphoribosylation of Nm to nicotinamide mononucleotide (NMN), followed by adenylylation. Judging from genome sequence data, the hyperthermophilic archaeon Thermococcus kodakarensis is supposed to utilize the four-step pathway, but the fact that the adenylyltransferase encoded by TK0067 recognizes both NMN and NaMN also raises the possibility of a two-step salvage mechanism. Here, we examined the substrate specificity of the recombinant TK1676 protein, annotated as nicotinic acid phosphoribosyltransferase. The TK1676 protein displayed significant activity toward Na and phosphoribosyl pyrophosphate (PRPP) and only trace activity with Nm and PRPP. We further performed genetic analyses on TK0218 (quinolinic acid phosphoribosyltransferase) and TK1650 (Nm deamidase), involved in de novo biosynthesis and four-step salvage of NAD + , respectively. The ΔTK0218 mutant cells displayed growth defects in a minimal synthetic medium, but growth was fully restored with the addition of Na or Nm. The ΔTK0218 ΔTK1650 mutant cells did not display growth in the minimal medium, and growth was restored with the addition of Na but not Nm. The enzymatic and genetic analyses strongly suggest that NAD + salvage in T. kodakarensis requires deamination of Nm and proceeds through the four-step pathway. IMPORTANCE Hyperthermophiles must constantly deal with increased degradation rates of their biomolecules due to their high growth temperatures. Here, we identified the pathway that regenerates NAD + from nicotinamide (Nm) in the

Renal transport and metabolism of nicotinic acid

International Nuclear Information System (INIS)

Schuette, S.; Rose, R.C.

1986-01-01

Renal metabolism and brush-border transport of nicotinic acid were studied in renal cortical slices and brush-border membrane vesicles exposed to a physiological concentration of vitamin (2.2-3.5 microM). Vesicle transport of [ 3 H]nicotinic acid was found to be Na+ dependent and concentrative. The presence of a Na+ gradient resulted in a fivefold increase in the rate of nicotinic acid uptake over that observed with mannitol and caused a transient nicotinic acid accumulation two- to fourfold above the equilibrium value. The effects of membrane potential, pH, and elimination of Na+-H+ exchange were also studied. Cortical slices and isolated tubules exposed to 2.2 microM [ 14 C]nicotinic acid took up vitamin and rapidly metabolized most of it to intermediates in the Preiss-Handler pathway for NAD biosynthesis; little free nicotinic acid was detectable intracellularly. The replacement of Na+ with Li+ in the bathing medium reduced total accumulation of 14 C label primarily as a result of reduced nicotinic acid uptake. Cortical tissue concentrated free nicotinic acid only when the involved metabolic pathways were saturated by levels of nicotinic acid far in excess of what occurs in vivo

Effects of Different Ratio of n-6/n-3 Polyunsaturated Fatty Acids on the PI3K/Akt Pathway in Rats with Reflux Esophagitis.

Science.gov (United States)

Zhuang, Jia-Yuan; Chen, Zhi-Yao; Zhang, Tao; Tang, Du-Peng; Jiang, Xiao-Yin; Zhuang, Ze-Hao

2017-01-30

BACKGROUND We designed this study to investigate the influence of different ratios of n-6/n-3 polyunsaturated fatty acid in the diet of reflux esophagitis (RE) rats' and the effect on the PI3K/Akt pathway. MATERIAL AND METHODS RE rats were randomly divided into a sham group and modeling groups of different concentrations of n-6/n-3 polyunsaturated fatty acid (PUFA): 12:1 group, 10:1 group, 5:1 group, and 1:1 group. RT-PCR and Western-blot were used to detect the expression of PI3K, Akt, p-Akt, NF-κBp50, and NF-κBp65 proteins in esophageal tissue. RESULTS In the n-6/n-3 PUFAs groups the expression of PI3K, Akt, p-Akt, nf-κbp50, and NF-κBp65 mRNA decreased with the decrease in n-6/n-3 ratios in the diet. The lowest expression of each indicator occurred in the 1:1 n-6/n-3 group compared with other n-6/n-3 groups, the difference was statistically significant (p<0.05). CONCLUSIONS The inhibition of n-3 PUFAs in the development of esophageal inflammation in rats with RE was attributed to the function of PI3K/Akt-NF-κB signaling pathway.

Immunohistochemical Studies of the Kynurenine Pathway in Morphea

OpenAIRE

Noakes, Rowland; Mellick, Nick

2013-01-01

Cutaneous sclerosis, resembling that seen in subcutaneous morphea, is a feature of eosinophilic fasciitis and eosinophilia-myalgia syndrome, two conditions in which the kynurenine pathway is known to be activated. To investigate the possibility of activation of the kynurenine pathway in morphea, skin biopsies were taken from involved and non-involved sites in a series of three patients with morphea. Immunohistochemical stains for quinolinic acid and indoleamine 2,3-dioxygenase (IDO) were perf...

Cloning, Expression Profiling and Functional Analysis of CnHMGS, a Gene Encoding 3-hydroxy-3-Methylglutaryl Coenzyme A Synthase from Chamaemelum nobile

Directory of Open Access Journals (Sweden)

Shuiyuan Cheng

2016-03-01

Full Text Available Roman chamomile (Chamaemelum nobile L. is renowned for its production of essential oils, which major components are sesquiterpenoids. As the important enzyme in the sesquiterpenoid biosynthesis pathway, 3-hydroxy-3-methylglutaryl coenzyme A synthase (HMGS catalyze the crucial step in the mevalonate pathway in plants. To isolate and identify the functional genes involved in the sesquiterpene biosynthesis of C. nobile L., a HMGS gene designated as CnHMGS (GenBank Accession No. KU529969 was cloned from C. nobile. The cDNA sequence of CnHMGS contained a 1377 bp open reading frame encoding a 458-amino-acid protein. The sequence of the CnHMGS protein was highly homologous to those of HMGS proteins from other plant species. Phylogenetic tree analysis revealed that CnHMGS clustered with the HMGS of Asteraceae in the dicotyledon clade. Further functional complementation of CnHMGS in the mutant yeast strain YSC6274 lacking HMGS activity demonstrated that the cloned CnHMGS cDNA encodes a functional HMGS. Transcript profile analysis indicated that CnHMGS was preferentially expressed in flowers and roots of C. nobile. The expression of CnHMGS could be upregulated by exogenous elicitors, including methyl jasmonate and salicylic acid, suggesting that CnHMGS was elicitor-responsive. The characterization and expression analysis of CnHMGS is helpful to understand the biosynthesis of sesquiterpenoid in C. nobile at the molecular level and also provides molecular wealth for the biotechnological improvement of this important medicinal plant.

Cloning, Expression Profiling and Functional Analysis of CnHMGS, a Gene Encoding 3-hydroxy-3-Methylglutaryl Coenzyme A Synthase from Chamaemelum nobile.

Science.gov (United States)

Cheng, Shuiyuan; Wang, Xiaohui; Xu, Feng; Chen, Qiangwen; Tao, Tingting; Lei, Jing; Zhang, Weiwei; Liao, Yongling; Chang, Jie; Li, Xingxiang

2016-03-08

Roman chamomile (Chamaemelum nobile L.) is renowned for its production of essential oils, which major components are sesquiterpenoids. As the important enzyme in the sesquiterpenoid biosynthesis pathway, 3-hydroxy-3-methylglutaryl coenzyme A synthase (HMGS) catalyze the crucial step in the mevalonate pathway in plants. To isolate and identify the functional genes involved in the sesquiterpene biosynthesis of C. nobile L., a HMGS gene designated as CnHMGS (GenBank Accession No. KU529969) was cloned from C. nobile. The cDNA sequence of CnHMGS contained a 1377 bp open reading frame encoding a 458-amino-acid protein. The sequence of the CnHMGS protein was highly homologous to those of HMGS proteins from other plant species. Phylogenetic tree analysis revealed that CnHMGS clustered with the HMGS of Asteraceae in the dicotyledon clade. Further functional complementation of CnHMGS in the mutant yeast strain YSC6274 lacking HMGS activity demonstrated that the cloned CnHMGS cDNA encodes a functional HMGS. Transcript profile analysis indicated that CnHMGS was preferentially expressed in flowers and roots of C. nobile. The expression of CnHMGS could be upregulated by exogenous elicitors, including methyl jasmonate and salicylic acid, suggesting that CnHMGS was elicitor-responsive. The characterization and expression analysis of CnHMGS is helpful to understand the biosynthesis of sesquiterpenoid in C. nobile at the molecular level and also provides molecular wealth for the biotechnological improvement of this important medicinal plant.